Plasmodium falciparum in Kenya: high prevalence of drug-resistance-associated polymorphisms in hospital admissions with severe malaria in an epidemic area.

During an epidemic of Plasmodium falciparum malaria in Chogoria, Kenya, P. falciparum DNA was collected from 24 cases of severe malaria admitted to hospital for parenteral quinine treatment. These patients had all failed first- (chloroquine) and second-line (sulfadoxine-pyrimethamine or amodiaquine) drug treatments. Twenty-two (92%) of the 24 patients sampled carried parasites with the (Asn)86(Tyr) point mutation in the pfmdr1 gene (chromosome 5), 20 (83%) had an (Asp)1246(Tyr) mutation and 18 (82%) had both of these mutations. These alleles are both reported to be associated with chloroquine-resistance. Polymorphisms in the cg2 gene (chromosome 7) are also associated with chloroquine resistance, and 18 (75%) of the 24 parasite samples each had the cg2 and pfmdr1 polymorphisms. These 18 samples also had the mutations associated with resistance to pyrimethamine and sulfadoxine: (Asn)51(Ile), (Cys)59(Arg) and (Ser)108(Asn) of gene dhfr (chromosome 4) and (Ala)437(Gly) and (Lys)540(Glu) of dhps (chromosome 8), respectively. Genotyping of the parasites from all 24 patients revealed extensive diversity in the sequences for the merozoite surface antigens (MSA-1 and MSA-2) and the glutamate-rich protein (GLURP) and indicated that each sample contained more than one parasite clone. Although samples from non-admitted malaria cases were not available, it appears that drug resistance may have played an important role in the development of severe malaria in this epidemic.

A comparative trial of standard or high-dose S subunit recombinant hepatitis B vaccine versus a vaccine containing S subunit, pre-S1, and pre-S2 particles for revaccination of healthy adult nonresponders.

The efficacy of 10-microg and 40-microg hepatitis B vaccines was compared with that of an investigational vaccine containing pre-S1, pre-S2, and S subunit particles (mixed particle vaccine) in inducing protective anti-hepatitis B surface antigen (anti-HBs) concentrations in 46 otherwise healthy persons who previously did not develop measurable levels of antibodies to at least one complete course of vaccine. A statistically significant difference was seen in the percentage of subjects who developed protective levels of anti-HBs (> or = 10 mIU/mL) with three 40-microg doses of S subunit vaccine versus the other groups. One hundred percent of the 40-microg dose group developed protective anti-HBs titers. No difference in adverse effects was noted.

IgG antibody to pneumococcal capsular polysaccharide in human immunodeficiency virus-infected subjects: persistence of antibody in responders, revaccination in nonresponders, and relationship of immunoglobulin allotype to response.

Human immunodeficiency virus (HIV)-infected persons are less likely than are noninfected persons to respond to vaccination with pneumococcal polysaccharides (PPS). Among those who respond, however, similar IgG levels may be achieved. HIV-infected men immunized with pneumococcal vaccine were classified as high- or low-level responders (IgG > or = 1 microgram/mL for > or = 3 of 5 PPS [high] or for < or = 1 PPS [low]). One and 2 years after immunization, geometric mean IgG levels and the percentages of subjects with IgG levels > or = 1 microgram/mL were similar for HIV-infected and for healthy high-level responders (controls) for all PPS except for serotype 8. Among HIV-infected low-level responders, revaccination with a double dose of pneumococcal vaccine did not stimulate IgG responses. Responsiveness of HIV-infected white patients was significantly associated with the Km(1)- negative allotype. These findings support current general recommended guidelines for administering pneumococcal vaccine to HIV-infected persons. Nonresponders will not benefit from revaccination.

Evidence that bismuth salts reduce invasion of epithelial cells by enteroinvasive bacteria.

The effects of sublethal concentrations of bismuth salts on bacterial invasion of mammalian cells were investigated. Pepto-Bismol, bismuth subsalicylate, and bismuth oxychloride, produced by interacting bismuth subsalicylate and simulated gastric juice, in suspension at concentrations as low as 1.4 mM significantly interfered with the invasion of RPMI-4788 cells by two different strains of Yersinia enterocolitica. Invasion of the mammalian epithelial cells by other enteric bacteria was also reduced significantly by some of these bismuth salts. Commercially obtained bismuth oxychloride, bismuth sulfide, and sodium salicylate had no affect on invasion by Y. enterocolitica. Exposure of Y. enterocolitica 8081c to Pepto-Bismol for as brief a time as 5 min was sufficient to produce the inhibitory effect. Removal of bismuth bound to bacteria by sodium potassium tartrate did not reverse the inhibition. Electron-dense deposits are observed in Y. enterocolitica 8081c exposed to bismuth subsalicylate, suggesting that interference of invasion may result from bismuth permeation of the bacterial cell wall.

Deposition of bismuth by Yersinia enterocolitica.

Yersinia enterocolitica 8081c cultures in exponential growth were incubated for 1 h in 0.1% microcrystalline bismuth subsalicylate (BSS) suspensions. Scanning electron microscopy (SEM) revealed microcrystals directly bound to BSS-treated bacteria. Energy dispersive spectroscopy (EDS) X-ray microanalysis of the attached microcrystals confirmed that the crystals were the microcrystalline BSS. X-ray spectra positive for bismuth were also obtained by SEM-EDS X-ray microanalysis of whole bacteria, suggesting metal incorporation into the bacteria in regions absent of bound microcrystals. Transmission electron microscopy of thin sections of embedded preparations of BSS-treated exponential-growth-phase bacteria showed electron-dense deposits in the periphery of the bacteria. Y. enterocolitica cultures that were in stationary phase at the time of incubation with microcrystalline BSS showed no evidence of the electron-dense deposits and EDS spectra were negative for bismuth. Bacteria incubated in the absence of microcrystalline BSS also lacked electron-dense deposits. Scanning transmission electron microscopy used in conjunction with EDS X-ray microanalysis to view and analyze semi-thick sections (250-300 nm) of embedded preparations of BSS-treated bacteria in exponential growth confirmed that the electron-dense deposits at the periphery of the bacteria are the sites of bismuth depositions.

140-MDa plasmids of Escherichia coli and Shigella flexneri 5 do not influence phagocytosis.

Enteroinvasive Escherichia coli (EIEC) and Shigella flexneri possessing a 140-megadalton (MDa) plasmid are capable of invading intestinal epithelial cells and causing dysentery. To determine if this plasmid affected phagocytosis of the organisms by leukocytes, we studied the in vitro phagocytosis of isogenic pairs of EIEC and S. flexneri 5 which differed only in the presence or absence of the 140-MDa plasmid. In addition five EIEC strains containing 140-MDa plasmids as well as one non-enteroinvasive E. coli strain possessing a 120-MDa plasmid were studied. The 140-MDa plasmid did not affect phagocytosis of these bacteria by normal human blood neutrophils or monocytes.

An in vitro model of Chlamydia trachomatis infection in the regenerative phase of the human endometrial cycle.

An in vitro model of the regenerative phase of the human endometrial cycle was developed in order to study the growth of Chlamydia trachomatis during the period following menses. Glandular epithelial fragments were prepared from curettings of endometria and explanted onto coated substrata. Epithelial cells migrated rapidly from the explant in a fashion which closely mimicked the regeneration of the surface epithelium after menses. The cultures were then experimentally infected with C. trachomatis serotype E at various times during formation of the outgrowth. Chlamydial inclusions developed both within the explants and in the outgrowing epithelial sheets. They were also found in isolated epithelial and non-epithelial cells. However, the most striking feature of chlamydial inclusion development within these cultures was the tendency for inclusions to be located in cells at the periphery of the epithelial sheets. This was partly due to the failure of the cells within the sheets to bind chlamydiae after centrifugation of the organisms onto the culture and partly due to a phenomenon similar to phagokinesis. During this process infectious chlamydial particles were cleared from the substratum by migrating cells with free motile edges, which occasionally led to internalization and inclusion development within these cells.

Phagocytes in cell suspensions of human colon mucosa.

Because little is known of the phagocytes of the human colon we enumerated these cells in mucosal suspensions and studied their phagocytic activity. Phagocyte rich suspensions were made by EDTA collagenase dissociation followed by elutriation centrifugation. Phagocytosis was evaluated by measuring cellular radioactivity after incubation of phagocytes with 3H-adenine labelled E coli ON2 and checked microscopically. Dissociation of normal mucosa from colorectal neoplasms yielded means of 1.9 X 10(6) eosinophils, 1.4 X 10(6) macrophages and 2 X 10(5) neutrophils per gram of mucosa. Visually normal mucosa of inflammatory states yielded 2.2 X 10(6) eosinophils, 2.3 X 10(6) macrophages and 7 X 10(5) neutrophils per gram of mucosa. Phagocyte rich suspensions of normal mucosa from tumour patients phagocytosed 21.8% of a pool of opsonised tritiated E coli ON2 and by microscopy 100% of mucosal neutrophils ingested bacteria, 83% of eosinophils were phagocytic, and 53% of macrophages contained bacteria. These results suggest that in the human colonic mucosa, the eosinophil is more abundant than the macrophage and the per cent of those cells exhibiting phagocytosis is intermediate between that of the macrophage and the neutrophil. Thus these three types of cells are actively phagocytic and share the potential for a major role in host defence against invasive enteric bacteria.

Eosinophils of human colonic mucosa: C3b and Fc gamma receptor expression and phagocytic capabilities.

Because little is known about eosinophils of the human intestine, we measured their C3b and Fc gamma receptor expression and phagocytic activity in mucosal suspensions from colon resections for large bowel neoplasms. Enzymatically dissociated suspensions were enriched for eosinophils by countercurrent centrifugation. C3b and Fc gamma receptors were measured by immunofluorescent assays with flow cytometry. Phagocytosis of Escherichia coli ON2 was determined by an in vitro microscopic method. Suspensions of normal tissue from neoplasm resections yielded 1.8 X 10(6) eosinophils/g mucosa, and these cells were more numerous than either macrophages or neutrophils. Fivefold enrichment was achieved by countercurrent centrifugation, and 75% of these cells expressed C3b receptors and 90% expressed Fc gamma receptors. Sixty-seven percent of mucosal eosinophils were phagocytic for E. coli ON2 and ingested a mean of 4.7 bacteria per cell. Eosinophils accounted for more overall phagocytic activity than either neutrophils or macrophages.

Pulmonary infections due to Legionella in immunocompromised patients.

At present, 11 different species of Legionella have been implicated in human disease. It has become apparent that disease caused by Legionella is acquired from a variety of environmental sources and that water is the factor that links many of them. Patients who are immunosuppressed, such as individuals receiving cancer chemotherapy or therapy designed to prevent organ rejection, are particularly susceptible to such environmental sources. It appears that intact cell-mediated immunity is more important in host defense than are adequate numbers of granulocytes or immunoglobulin concentrations. Diagnostic steps should be undertaken in all patients developing nosocomial pneumonia who present with a picture suspicious for this disorder. In the meantime, appropriate antimicrobial therapy with erythromycin and rifampin should be begun. If clusters of cases are detected in a hospital, immediate steps should be taken to attempt to isolate the organism from any aqueous environmental sources, and if found appropriate, steps taken. Awareness of the threat of legionnaires' disease must be maintained among clinicians and hospital epidemiologists because it is unlikely that the problem of nosocomial legionnaires' disease will disappear.

Lack of association between genital mycoplasmas and infertility.

We studied the relation between colonization with Mycoplasma hominis and Ureaplasma urealyticum, and the results of infertility studies in 205 women with involuntary infertility of at least one year's duration. Isolation of M. hominis (but not of U. urealyticum) was significantly (P = 0.002) more common in patients with a history of pelvic inflammatory disease. However, no relation could be shown between these genital mycoplasmas and any of the following: evidence of prior pelvic inflammatory disease as determined by hysterosalpingography and laparoscopy; cervical inflammation; numbers and motility of spermatozoa on postcoital test; pyosemia; quality of cervical mucus; whether the cause of infertility was related to male or female factors, both, or neither; and occurrence and outcome of subsequent pregnancy. Mycoplasmas were cultured from only 10 of 203 endometrial biopsy specimens (4.9 per cent), and in no instance was inflammation associated with this finding. Out studies do not support a role for genital mycoplasmas in the cause of infertility.

Patterns of adnexal inflammatory damage: Chlamydia, the intrauterine device, and history of pelvic inflammatory disease.

In a study of 204 consecutive infertile couples, 58 women with adnexal abnormalities consistent with prior pelvic infection were identified. The status of those 58 subjects with respect to prior pelvic infection, prior intrauterine device use, and serologic evidence of past chlamydial infection was correlated with the types of adnexal abnormalities identified. Women with serologic evidence of past chlamydial infection were more likely to exhibit severe adhesions and hydrosalpinx formation, and hydrosalpinx formation was related to a history of clinically detected pelvic infection.

The kinetics of early inflammatory events during experimental pneumonia due to Legionella pneumophila in guinea pigs.

An animal model of Legionella pneumophila pneumonia was developed to study aerosol infection, pathogenesis, and pulmonary host defense mechanisms. Guinea pigs were exposed in an inhalation facility that limited the aerosol of L pneumophila to the snout. Bronchoalveolar lavage was used to sample airspace cells, secretions, and bacteria during developing infection in 79 exposed animals and 13 uninfected controls. An influx of polymorphonuclear neutrophils followed exponential bacterial growth during the initial three days of infection and coincided with limitation of the increase in bacteria recovered. A macrophage influx occurred at three to five days. Bacteria were eliminated from the lung by 11 days after exposure. Albumin in lavage fluid peaked at two days. Most viable L pneumophila organisms were associated with alveolar macrophages, whereas most of the bacteria associated with polymorphonuclear neutrophils were nonviable. Recruited, and possibly immune, defenses appear to be required for successful resolution of legionella pneumonia.

Activity of trimethoprim (TMP), sulphamethoxazole (SMX) and the combination against Bacteroides fragilis group isolates.

The in-vitro efficacy of trimethoprim/sulphamethoxazole (TMP/SMX) was tested against 59 isolates belonging to the Bacteroides fragilis group of bacteria and was shown to be dependent upon the inoculum size. With an inoculum of 3 X 10(5) colony forming units (cfu), 98% of these isolates were susceptible to the combination, whereas with a higher inoculum of 10(6) cfu, 88% were susceptible. None of the isolates were susceptible to less than 2 mg/l of TMP, whereas 57 (97%) were susceptible to SMX at the lower inoculum and 21 (36%) at the higher inoculum. These data indicate that TMP/SMX has moderate activity against organisms of the Bact. fragilis group when tested at an inoculum of 10(6) cfu.

Protein and antibody in lavage fluid of guinea pigs with Legionella pneumophila pneumonia.

Guinea pigs were infected in an inhalation facility that limited an aerosol of L. pneumophila to the snout, as previously reported in detail (Davis et al., 1982). Individual animals were sacrificed for study either immediately after exposure, at 16 hours, at days one through seven, or at 11 days. Bronchoalveolar lavage was carried out to obtain fluid to study the following: total protein, albumin and immunoglobulin G (IgG) concentrations, and the titer of antibody to L. pneumophila. Antibody also was measured in serum obtained at the time of sacrifice. Concentrations of total protein, albumin, and IgG in lavage fluids peaked 2 days after exposure and correlated with the appearance of maximal numbers of polymorphonuclear cells in the lungs. Presumably, this increased protein resulted from exudation of serum across the alveolar-capillary membrane, which loses its integrity secondary to pneumonia. However, the ratio of IgG/albumin was elevated in animals studied 11 days after exposure even though the concentration of albumin was normal by this time. One possible explanation for this observation is that IgG was being produced in the lung. Antibody in lavage fluid was detected 7, and 11 days post-exposure, and might be important in the recovery of guinea pigs from this infection.