Does examination of urinary sediment identify individuals with Gulf War syndrome? A pilot study.

BACKGROUND: Many veterans who were involved in the Persian Gulf theater of operations have had a variety of unexplained physical complaints, collectively called the Gulf War syndrome or similar names. There has been much debate on the issue and numerous publications, both in the medical and the lay press. A method for examining urinary sediment that was developed in an effort to identify nonculturable bacteria has been used in Gulf War veterans and was the basis for intensive antimicrobial therapy in many of them. METHODS: We evaluated eight Gulf War veterans with complaints compatible with Gulf War syndrome. Subjects were from various parts of the United States. A detailed history and physical examination were performed. Urine was obtained before and after prostatic massage (men) or before and after pelvic examinations (women) and was tested by a previously described microscopic method as well as by culture and conventional Gram stain. Age- and sex-matched healthy control subjects were tested similarly and concurrently. RESULTS: Two female Gulf War veterans had findings of Candida albicans and Klebsiella pneumoniae by conventional culture. The same organism types were seen both by the special method and by conventional Gram stain. All other subjects and controls were completely indistinguishable. CONCLUSION: Examining the urinary sediment by this elaborate method does not differentiate persons with Gulf War syndrome from normal, healthy control subjects who were never in the Persian Gulf area.

Algaemia due to Prototheca wickerhamii in a patient with myasthenia gravis.

Prototheca wickerhamii is a rare cause of systemic infection in humans. While some cases occur in previously healthy individuals, others are associated with a variety of preexisting diseases. Here we present, for the first time, a case of P. wickerhamii algaemia in a patient with myasthenia gravis. The patient was successfully treated with amphotericin B.

Inhibitory effect of 0 degree C storage on the proliferation of Yersinia enterocolitica in donated blood.

BACKGROUND: Yersinia enterocolitica is frequently identified in cases of bacterial sepsis due to red cell transfusion. One of the features that makes Y. enterocolitica particularly dangerous is that, unlike most other bacterial contaminants of blood components, this organism can actively multiply in currently recommended refrigerator temperatures (1-6 degrees C). The effect of a colder than normal storage temperature on Y. enterocolitica growth was investigated to determine whether bacteria growth could be reduced or inhibited at 0 degree C. STUDY DESIGN AND METHODS: Twenty-four units of freshly collected donated blood were obtained. Three sets of 7 units each were inoculated with Y. enterocolitica O:3, Y. enterocolitica O:20, and Y. enterocolitica O:5, 27, respectively. The remaining 3 units served as uninoculated controls. Each of the 24 bags was split into two equal aliquots, with one aliquot stored at 4 degrees C and the other at 0 degree C. Bacteria growth was measured twice weekly for 6 weeks. Endotoxin and hemoglobin levels were also measured at selected intervals. RESULTS: Bacteria growth was detected earlier and in higher concentrations in the aliquots stored at 4 degrees C. Twenty-two of the 42 inoculated aliquots had measureable bacteria growth. Thirteen aliquots had been maintained at 4 degrees C, and nine had been stored at 0 degree C. Sixteen of these 22 aliquots were matched pairs. Exponential growth was detected after 14 to 32 days in the 4 degrees C aliquots and after 28 to 39 days in the 0 degree C aliquots. Final bacteria counts were much higher in the 4 degrees C aliquots (10(5)-10(14) colony-forming units/mL) than in the 0 degree C aliquots (10(1)-10(4) colony-forming units/mL) on Day 42. Endotoxin was present in all 13 of the 4 degrees C aliquots with actively growing Y. enterocolitica. CONCLUSION: Storage of red cells at 0 degree C markedly prolongs the time required for Y. enterocolitica to achieve exponential grwoth and results in lower concentrations of bacteria.

The family pet as an unlikely source of group A beta-hemolytic streptococcal infection in humans.

This study examines the possibility of the family pet serving as a reservoir for group A beta-hemolytic streptococcal infections in humans. We obtained oropharyngeal cultures from children with acute pharyngitis and concurrent oropharyngeal cultures from their household pets. Children with culture-proved group A beta-hemolytic streptococcal pharyngitis were detected in 26 of 42 households surveyed. Oropharyngeal cultures were also collected from a group of children without pharyngitis and their pets. Additionally 149 dogs and cats from a local veterinary hospital were cultured from 371 body sites including the oropharynx, axilla and vagina. All beta-hemolytic bacterial isolates were identified by colonial and microscopic morphology, catalase and pyrrolidonylarylamidase production, bacitracin susceptibility and serogrouping. No group A beta-hemolytic streptococci were recovered from any of the body sites surveyed from a total of 230 animals. Based on these findings, the family pet seems to be an unlikely reservoir for group A beta-hemolytic streptococci.

Evaluation of Gen-Probe's Histoplasma capsulatum and Cryptococcus neoformans AccuProbes.

Gen-Probe's DNA probes were evaluated for use in the identification of clinical isolates of Histoplasma capsulatum var. capsulatum and Cryptococcus neoformans. Ninety-five mould-phase fungi were probed, including 41 isolates of H. capsulatum var. capsulatum. Similarly, 98 yeasts, including 42 C. neoformans isolates, were examined by using the C. neoformans DNA probe. In the study, both probes demonstrated 100% specificity and 100% sensitivity. Their use in the clinical laboratory may significantly reduce the time required for definitive identification of fungi.

Apophysomyces elegans as an agent of zygomycosis in a patient following trauma.

Apophysomyces elegans was isolated from the subcutaneous tissue, muscle and bone of a patient who had fallen from a height of 55 feet. Broad, sparsely septate hyphae were present in the tissue. Surgical debridement of wounds and amphotericin B treatment were not sufficient in controlling rapid tissue necrosis. Ultimately, amputation of the two affected limbs was necessary.

Achromobacter xylosoxidans. An unusual neonatal pathogen.

Perinatal acquisition of a rare pediatric pathogen, Achromobacter xylosoxidans, with evidence for in utero transmission, is described. Cultures from the mother and neonate demonstrated A. xylosoxidans. An ascending bacterial infection in the mother with clinical chorioamnionitis is presented as the probable mode of transmission. Postmortem examination of the infant confirmed Achromobacter meningitis. In contrast to the current case with transmission from mother to neonate, previously published neonatal cases of Achromobacter infections indicate that nosocomial transmission of the organism is most common (79%). In addition, the literature review revealed a high mortality associated with meningitis (77%), frequent hydrocephalus, and subsequent neurologic sequelae (36%). To the authors' knowledge, this is the first documented case of maternal-fetal transfer of A. xylosoxidans.

Herpes simplex virus type 2 meningoencephalitis resistant to acyclovir in a patient with AIDS.

A case is reported of relapsing fatal meningoencephalitis caused by a neurovirulent thymidine kinase-positive (TK+) type 2 herpes simplex virus (HSV) that developed thymidine kinase deficiency (TK-) during intravenous acyclovir therapy. A patient with AIDS was admitted for acyclovir treatment of a persistent perirectal herpetic ulcer. He subsequently developed meningoencephalitis. A TK+ type 2 HSV was isolated from a brain biopsy specimen. A progressive and fatal relapse occurred, and a TK- type 2 HSV was isolated from his cerebrospinal fluid. Restriction endonuclease analysis of viral DNA from perianal, brain, and cerebrospinal fluid isolates were similar, suggesting that they were the same viral strain. Animal virulence studies indicated significant cutaneous virulence in immunocompromised mice models for the TK- isolates. This case is notable because TK- HSV have, in the past, lacked neurovirulence and because acyclovir resistance developed during therapy and caused the patient's death.

Detection of piluslike structures on clinical and environmental isolates of Vibrio vulnificus.

Twenty clinical isolates of Vibrio vulnificus were compared with 10 environmental strains by using electron microscopy and agglutination assays with human erythrocytes, guinea pig erythrocytes, and Saccharomyces cerevisiae. In addition, the isolates were tested for ability to adhere to the human epithelial cell lines HEp-2 and A549. When examined by electron microscopy, 16 (80%) of the 20 clinical isolates demonstrated the presence of piluslike structures; the composition of the bacterial populations ranged from 0 to 68% piliated cells. In contrast, only 3 (30%) of the 10 environmental isolates were piliated, with a range from 0 to 16% piliated cells. A significant association between the presence of piliated cells and the isolate source was found (P less than 0.05). None of the 30 strains agglutinated erythrocytes or yeast cells. V. vulnificus adherence results obtained with HEp-2 cells showed 10 (50%) of 20 clinical isolates and 0 (0%) of 10 environmental isolates with averages of greater than 10 adherent bacteria per cell, demonstrating a correlation between attachment and the isolate source (P less than 0.05). Selected strains were tested to determine whether methyl alpha-D-mannopyranoside, fructose, or alpha-L-(-)-fucose would inhibit bacterial adherence to HEp-2 cells. Multiple patterns of adherence inhibition were observed. Adherence to A549 cells showed 8 (40%) of 20 clinical isolates and 0 (0%) of 10 environmental strains with averages of greater than 10 adherent bacteria per cell. A statistical association between attachment and the isolate source was demonstrated (P less than 0.05). These data suggest that the presence of piluslike structures and the ability to adhere to human epithelial cell lines may be more closely associated with V. vulnificus isolates from clinical specimens than with environmental strains.

Modified spin-amplified adsorption procedure with conventional tissue culture tubes for rapid detection and increased recovery of herpes simplex virus from clinical specimens.

Conventional culture tubes were used in a modification of the spin-amplified adsorption procedure for recovery of herpes simplex virus (HSV) from clinical specimens. The sensitivity of isolation of HSV from 864 specimens adsorbed by the spin-amplified method was 100% (127 of 127), compared with 88.2% (112 of 127) for stationary-phase-adsorbed specimens. Cytopathic effect developed more rapidly in 32.1% (36 of 112) of isolates adsorbed by spin amplification than in those adsorbed by stationary means. In a separate quantitative study, cultures of HSV type 1 adsorbed by spin amplification yielded higher antigen levels and greater cytopathic effect than stationary-phase-adsorbed cultures. Cells grown in conventional tissue culture tubes may be used in a spin-amplified adsorption for rapid detection and increased sensitivity of HSV isolation.

Multiple drug-resistance in Shigella flexneri isolated from a patient with human immunodeficiency virus.

Persistent shigellosis, due to Shigella flexneri resistant to multiple antibiotics, developed in a 40-yr-old homosexual man with human immunodeficiency virus infection. The Shigella strain demonstrated resistance to ampicillin, tetracycline, and trimethoprim-sulfamethoxazole. Although Shigella flexneri isolates resistant to trimethoprim-sulfamethoxazole are uncommon in the United States, laboratories should monitor resistance patterns through routine in vitro susceptibility testing.

Distribution of type 1 and P pili on uropathogenic Escherichia coli O6.

The distribution of type 1 and P pili on individual cells of an O6 uropathogenic Escherichia coli strain, 6260, was determined immunologically with pilus-specific monoclonal antibodies by indirect immunofluorescence and immunogold electron microscopy. Variations in pilus expression under different culture conditions were monitored with an indirect immunofluorescence assay; 63% of piliated cells expressed type 1 pili when grown on agar at 37 degrees C versus 14 to 38% when grown in broth at 37 degrees C. In contrast, generally fewer cells with P pili (18 to 44%) were detected on agar than when grown in broth (up to 86%). Both type 1 and P pili were absent from cells cultured at 20 degrees C. Immunogold and immunofluorescence double labeling techniques with monoclonal antibodies 11-2 and 91-1 were used to study subpopulations of cells with type 1 and P pili; 39 to 41% of the piliated cells demonstrated only type 1 pili, and 12 to 16% of the cells showed only P pili. The immunogold method proved more sensitive than the immunofluorescence technique for detecting subpopulations expressing both pili types simultaneously, 19 versus 7%. We observed variations between type 1 and P pili, both in expression on individual cells and in the distribution of subpopulations of cells.

Fimbrial phase variation and DNA rearrangements in uropathogenic isolates of Escherichia coli.

Having previously shown that the oscillating on-off expression (phase variation) of type 1 fimbriae in Escherichia coli is regulated genetically by an invertible element of DNA, we wished to determine whether E. coli isolates recovered from infected humans behaved in similar fashion. We examined four different clinical isolates that expressed type 1 fimbriae, P fimbriae, or both. Using, in Southern blot analysis, a DNA probe from the type 1 fimbrial switch, that hybridized to one DNA band from phase-on bacteria and to two DNA bands from phase-off bacteria, we found that the three clinical isolates expressing type 1 fimbriae contained the same invertible switch previously seen in the K-12 isolate. Employing a similar approach to characterize the on-off expression of P fimbriae, we used a DNA probe containing the known transcriptional signals for P fimbriae. Although we detected DNA rearrangement in the two strains expressing P fimbriae, unlike the case for type 1 fimbriae, the rearrangement did not correlate with the on-off state of the P fimbriae. Rather, the DNA rearrangement correlated with the environmental conditions of growth of the bacteria from which the DNA was isolated. These results confirm the notion that P fimbriae expression and type 1 fimbriae expression are controlled differently.

Utilization of anion-exchange chromatography and monoclonal antibodies to characterize multiple pilus types on a uropathogenic Escherichia coli O6 isolate.

Multiple pilus types from a uropathogenic strain of Escherichia coli O6, strain 6260, were characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), high-pressure liquid chromatography, binding assays, and erythrocyte adsorption. In addition, monoclonal antibodies were raised against purified pili of E. coli 6260 and used for immunological characterization. SDS-PAGE analysis of the purified pili showed at least three different subunits with molecular weights of 15,700, 17,800, and 19,300. SDS-PAGE analysis of four protein peaks from anion-exchange chromatography of intact pili showed polypeptides with molecular weights of 19,300 (fraction 1), 15,700 (fraction 2), and 17,800 and 15,700 (both fractions 3 and 4). Erythrocyte adsorption of the whole-pilus preparation removed the 17,800-molecular-weight subunit (17.8K subunit) and reduced the 15.7K subunit. Pili from an isogenic hemagglutination-negative variant of E. coli 6260, showing only the 15.7K and 19.3K subunits by SDS-PAGE, lacked the 17.8K subunit of fractions 3 and 4 present in the parent high-pressure liquid chromatography profile. Our data suggest that two of the pilus subunits, the 15.7K and 17.8K subunits, mediate mannose-resistant agglutination of human erythrocytes. Pili in fractions 1 and 2 from the parent strain bound specifically to mannose residues, while pili in fraction 4 bound to P-coated horse erythrocytes; no receptor specificity was identified for pili in fraction 3. Immunological analysis by the immunoblot technique showed that monoclonal antibody 11-2 reacted with the 19.3K subunit, monoclonal antibodies 34-3 and 73-3 reacted with the 15.7K subunit, and monoclonal antibodies 81-1, 82-1, and 91-1 reacted with polymers of subunits retained in the stacking gel. Intact pili precipitated by any of the six monoclonal antibodies showed two polypeptides by SDS-PAGE: 15.7K and 19.3K polypeptides for monoclonal antibody 11-2, and 15.7K and 17.8K polypeptides for monoclonal antibodies 34-3, 73-3, 81-1, 82-1, and 91-1. The cross-reactivity of the monoclonal antibodies with purified pili from other E. coli strains was determined by enzyme-linked immunosorbent assay. Monoclonal antibody 11-2 showed no significant cross-reactivity with heterogeneous pili. In contrast, the other monoclonal antibodies showed equivalent or greater reactivity with P pili from heterologous strains as compared with reactivity with E. coli 6260 pili.(ABSTRACT TRUNCATED AT 400 WORDS)

Mannose-resistant hemagglutination and P receptor recognition of uropathogenic Escherichia coli isolated from adult patients.

Adhesions of 211 strains of uropathogenic Escherichia coli and 19 strains of normal fecal E. coli were characterized by patterns of agglutination with human erythrocytes, Saccharomyces cerevisiae, and horse erythrocytes coated with the P blood-group receptor (P). Mannose-resistant (MR) hemagglutination was significantly associated with P agglutination (P less than .001). E. coli expressing MR and/or P (MR/P) agglutinins concurrently with mannose-sensitive (MS) agglutinins predominated in all clinical categories. The highest percentage of E. coli demonstrating MR/P agglutinins, in the absence of MS agglutinins, was recovered from patients with acute pyelonephritis (35%) compared with percentages of patients with chronic pyelonephritis (13%), asymptomatic bacteriuria (16%), cystitis (11%), and normal fecal control E. coli (11%). Sixty-nine percent of E. coli isolates causing acute pyelonephritis agglutinated P-coated horse erythrocytes compared with only 11% of the fecal isolates. Strains expressing MR/P agglutinins (in the absence of MS agglutinins) isolated from patients with acute pyelonephritis, chronic pyelonephritis, and asymptomatic bacteriuria were significantly associated with the presence of antibody-coated bacteria in patients' urine sediments (P less than .010), an observation indicative of an immune response associated with bacterial invasion of host tissues.