bluuebunny/crossref_metadata_2025 huggingface dataset:
Iron-vibriobactin transport system is not required for virulence of Vibrio cholerae
Authors: Sigel S P , Stoebner J A , Payne S M
Published: January 2020

The possible requirement of a functional siderophore (vibriobactin)-mediated iron transport system in the pathogenicity of Vibrio cholerae was determined. Two mutants of V. cholerae defective in the iron-vibriobactin transport system were examined for their ability to multiply and elicit diarrhea in infant mice. One mutant, 40130, was unable to synthesize vibriobactin. The second mutant, 1510, was able to synthesize, but not transport, the siderophore. Both mutants retained the ability to multiply and produce disease in the infant mouse, and virulence was indistinguishable from the parent strains. This indicates that a functional iron-vibriobactin transport system is not essential for cholera pathogenesis. These mutants, like the wild-type strains, were found to have a ferric citrate iron uptake system and could utilize citrate or asparagine for growth in low-iron medium. Compounds of this type may increase the availability of iron to V. cholerae in the host.
Cell cycle distribution patterns and generation times of L929 fibroblast cells persistently infected with Coxiella burnetii
Authors: Baca O G , Scott T O , Akporiaye E T , DeBlassie R , Crissman H A
Published: January 2020

Coxiella burnetii established a persistent infection of various cell lines including L929 mouse fibroblasts. Although the basis for such persistence is unknown, the phenomenon does require continual growth, proliferation, and maintenance of viability of the host cells. We examined the effect of short- and long-term infection on the host cell's generation time and cell cycle. Flow cytometric studies of actively growing normal and infected cells stained with mithramycin or propidium iodide revealed no significant difference in cell cycle distribution patterns or changes in ploidy level associated with persistent infection with either phase I or phase II C. burnetii. The population doubling times of infected and normal cells were similar.
Physical and genetic analysis of DNA regions encoding the immunoglobulin A proteases of different specificities produced by Haemophilus influenzae
Authors: Bricker J , Mulks M , Moxon E R , Plaut A G , Wright A
Published: January 2020

The structural gene for immunoglobulin A protease (iga) from Haemophilus influenzae serotype d was cloned in pBR322. The gene was used as a probe for Southern hybridization analysis of chromosomal DNA from the five other H. influenzae serotypes (a, b, c, e, and f). In most cases strains from a single serotype exhibited a distinct pattern of restriction fragment(s) homologous to the iga gene probe which was unique for that serotype. Serotype f strains were unique in that they gave two distinct patterns of homologous restriction fragments which correlated well with the production of two different protease types by members of this group. An iga mutant of H. influenzae serotype d was isolated by introducing a 4-base-pair insertion into the cloned iga gene and using the altered DNA for transformation of an H. influenzae recipient. The resulting iga- mutant produced no immunoglobulin A protease but was otherwise indistinguishable from its iga+ parent in growth characteristics. Transformation of mutant cells with chromosomal DNA isolated from either a serotype d or a serotype c strain gave rise to iga+ transformants. Those obtained with serotype d DNA produced a type 1 protease, whereas those obtained with serotype c DNA produced either a type 1 protease (characteristic of serotype d) or a type 2 protease (characteristic of serotype c). Southern analysis of the latter transformants, using the iga gene probe, indicated that the type 1 transformants had a serotype d pattern of restriction fragments whereas the type 2 transformants had either a serotype c or a novel pattern of restriction fragments. These results indicate that there is considerable homology between the iga genes of the various serotypes and that the homologous sequences identified with the serotype d probe are the immunoglobulin A protease-coding sequences in each case.
Increased sensitivity of Corynebacterium parvum-treated mice to toxic effects of indomethacin and lipopolysaccharide
Authors: Hart D A
Published: January 2020

Female BALB/c and C3H/HeJ mice develop increased sensitivity to the toxic effects of indomethacin after injection of nonviable Corynebacterium parvum. The increased sensitivity developed within 4 days of intraperitoneal injection of the organisms and started to resolve 14 days after injection. The development of increased sensitivity was dependent on the quantity of organisms injected and the concentration of indomethacin utilized. The effect was not observed when C. parvum-treated animals were injected with aspirin. C. parvum-treated BALB/c mice also developed increased sensitivity to E. coli lipopolysaccharide (LPS). Although increased sensitivity to LPS and indomethacin paralleled each other in BALB/c mice, the experiments with the LPS-resistant C3H/HeJ mice indicated that the two phenomena could be separated. The pyridine extract residue of C. parvum was as effective as C. parvum whole cells in inducing indomethacin and LPS sensitivity. Therefore, activation of the reticuloendothelial system is probably a critical element in the induction of sensitivity to these agents.
Antibodies to Bordetella pertussis in human colostrum and their protective activity against aerosol infection of mice
Authors: Oda M , Cowell J L , Burstyn D G , Thaib S , Manclark C R
Published: January 2020

Colostrum samples from Indonesian mothers were assayed for antibodies which agglutinate Bordetella pertussis and for antibodies to the filamentous hemagglutinin and the lymphocytosis-promoting factor of B. pertussis. Agglutinins were assayed by a microtiter method, and 36 of 58 samples tested (62%) had titers above 1:10 (range, less than 1:10 to 1:160). An enzyme-linked immunosorbent assay detected anti-filamentous hemagglutinin in 39 of 60 samples (65%) and anti-lymphocytosis-promoting factor in 26 of 60 samples assayed (43%). A total of 52 samples (87%) were positive for at least one of these antibodies. Pooled colostrum samples were separated by affinity chromatography into fractions enriched secretory immunoglobulin A (sIgA) or IgG and examined for their ability to passively protect suckling mice from aerosol challenge with B. pertussis. Samples (160 micrograms of protein) were given intraperitoneally 90 min before challenge. Death, rate of gain in body weight, and leukocytosis were used as indicators of illness. Colostrum containing anti-lymphocytosis-promoting factor or agglutinins was protective, whereas colostrum lacking these but containing anti-filamentous hemagglutinin gave little protection. The sIgA-enriched and IgG-enriched fractions appeared to be equal in their ability to protect against respiratory challenge with B. pertussis.
Prevention of adhesion by indigenous bacteria to rabbit cecum epithelium by a barrier of microvesicles
Authors: Hill R H
Published: January 2020

None of 74 strains of anaerobic bacteria cultured from the mucosal epithelial interface of the rabbit cecum adhered to isolated brush borders in experiments in which enteropathogenic Escherichia coli RDEC-1 was used as an adherent control. Scanning electron microscopy of the surfaces of the cecal epithelia confirmed that few organisms were in contact with the villus surface, but transmission electron microscopy revealed a layer of microvesicles up to 50 nm in diameter between the microvilli and mucous gel. The evidence indicates that there is no significant epithelium-adherent flora in the cecum but that a microvesicular layer could contribute significantly to prevention of adherence by both normal and potentially pathogenic bacteria.
In vitro evidence that human airway lysozyme is cleaved and inactivated by Pseudomonas aeruginosa elastase and not by human leukocyte elastase
Authors: Jacquot J , Tournier J M , Puchelle E
Published: January 2020

The in vitro effects of Pseudomonas aeruginosa elastase (P. aeruginosa E) and of human leukocyte elastase on human airway lysozyme (HAL) were investigated. P. aeruginosa E inactivated and cleaved HAL, whereas human leukocyte elastase had no effect. Total inactivation of HAL by P. aeruginosa E was observed after 120 min of incubation at 37 degrees C, for an elastase-to-lysozyme molar ratio of 1:5. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of reaction mixtures containing HAL and P. aeruginosa E in an elastase-to-lysozyme molar ratio of 1:10 showed a progressive disappearance of the HAL band upon increasing the incubation time with P. aeruginosa E. Gel filtration chromatography indicated that HAL was cleaved into at least three peptide fragments. The cleavage of HAL by P. aeruginosa E was accompanied by parallel losses of its bacteriolytic activity and its immunoreactive property.
Model of experimental chronic osteomyelitis in rats
Authors: Rissing J P , Buxton T B , Weinstein R S , Shockley R K
Published: January 2020

We describe here a Sprague-Dawley rat model for chronic osteomyelitis. Staphylococcus aureus and sodium morrhuate were implanted by either microdrilling or direct needle injection into the tibiae of rats. Of 107 rats, 87 (81%) developed osteomyelitis when a high-speed drill was used for implantation, and 27 (51%) of 53 rats developed osteomyelitis by direct needle inoculation (chi square = 9.81, P less than 0.01). Demonstrated histopathological changes included the presence of resorption bays filled with osteoclasts. Quantitative microbiological monitoring of tibial count confirmed disease chronicity, yielding stable numbers of CFU (10(6.29 +/- 0.27) ) of S. aureus over 70 days. Infected animals became anemic and lost weight. The erythrocyte sedimentation rates and leukocyte counts were not elevated. Roentgenograms provided the best correlation with the number of organisms in infected tibiae (r2 = 0.80). Rats with infected tibiae were treated with either oxacillin (120 mg/kg per day) or ceftriaxone (50 mg/kg per day). Treatment over 14 or 28 days reduced S. aureus counts in tibiae but did not reliably sterilize infected bones, suggesting that this model was resistant to prolonged antimicrobial therapy.
Resistance to adenovirus infection after administration of Bordetella pertussis vaccine in mice
Authors: Winters A L , Baggett D W , Benjamin W R , Brown H K , Klein T W
Published: January 2020

Treatment of mice with Bordetella pertussis vaccine rendered mice resistant to mouse adenovirus infection. The resistant state took at least 5 days to develop, and susceptibility returned to a portion of the test population 35 days after treatment. Transient resistance developed in congenitally athymic mice also. Treatment with a dose of 25 micrograms (dry weight) of B. pertussis vaccine protected approximately 50% of the test population. Vaccines prepared from several different strains of B. pertussis were capable of inducing resistance, and the induction of resistance was not dependent on the mouse strain used for testing. Cross-reacting antibodies capable of neutralizing the virus or protecting against a challenging infection were not induced by treatment with B. pertussis vaccine.
Cholate-dependent killing of Giardia lamblia by human milk
Authors: Gillin F D , Reiner D S , Gault M J
Published: January 2020

We showed previously that nonimmune human milk (NHM) kills Giardia lamblia trophozoites in vitro and presented evidence that killing requires the bile salt-stimulated lipase of milk. Since this enzyme is activated by bile salts, killing should be dependent on the presence of bile salts. We now show that killing by fresh NHM or NHM stored at -70 degrees C is totally dependent on sodium cholate (a bile salt). With less than 0.4 mM cholate, no parasites were killed, whereas with 1 mM cholate, greater than 99.7% were killed by 5% NHM in 30 min. Moreover, killing activity was completely heat labile. The G. lamblia-killing activity of human milk was greatly altered by storage at -10 or -20 degrees C. In less than 23 days, the 50% lethal dose decreased, cholate dependence was lost, and killing activity became heat stable. In contrast, the activity of milk stored at -70 degrees C remained unchanged. Milk lipase activity, like killing activity, became cholate independent during storage at -10 or -20 degrees C. On the basis of these results, we hypothesize that killing of G. lamblia by fresh NHM or NHM stored at -70 degrees C depends on bile salt-stimulated lipase, which must be activated by bile salts. In contrast, NHM stored at -20 degrees C accumulated free fatty acids which kill G. lamblia. In support of this thesis, milk stored at -10 degrees C had a concentration of 18.7 mM free fatty acids compared with only 1.1 mM in an identical sample stored at -70 degrees C.
Invasion of HEp-2 cells by fecal isolates of Aeromonas hydrophila
Authors: Lawson M A , Burke V , Chang B J
Published: January 2020

Human diarrheal isolates of Aeromonas hydrophila were found to be invasive in HEp-2 cell monolayers. Microscopic examination of two isolates from patients with symptoms of dysentery revealed that infected HEp-2 cells contained up to 50 bacteria localized within areas of the cytoplasm. A quantitative invasion assay was developed, using viable counts of total intracellular bacteria. Five fecal isolates of A. hydrophila were classified as invasive, with an average of 0.1 up to 2 bacteria per HEp-2 cell, as compared to 7 for an invasive Escherichia coli strain. 3 other fecal isolates, 1 reference strain, and 10 water isolates of A. hydrophila were similar to a noninvasive E. coli strain, with less than 0.008 bacteria per HEp-2 cell. All isolates were screened for plasmid DNA; no common plasmid was found in the invasive strains, and the loss of a 6.0-megadalton plasmid from one of these strains had no observable effect on invasiveness. Thus some A. hydrophila isolates are capable of cellular penetration and replication, and this may be an important chromosomally determined virulence property of the organism.
Cloning the gene for Congo red binding in Shigella flexneri
Authors: Daskaleros P A , Payne S M
Published: January 2020

The ability to bind the dye Congo red from agar medium is associated with virulence of Shigella species. DNA sequences conferring this property have been cloned from a large, 140-kilobase plasmid of Shigella flexneri into a plasmid vector. This recombinant plasmid does not fully restore virulence to S. flexneri isolates which have lost the large plasmid. This indicates that other genes present on the 140-kilobase plasmid must also be required for virulence of S. flexneri. The cloned fragment contains a copy of the insertion sequence IS1 closely linked to the gene for Congo red binding.
Modulation of pulmonary clearance of bacteria by antioxidants
Authors: Pesanti E L , Nugent K M
Published: January 2020

To further delineate the mechanisms underlying murine pulmonary defenses against bacterial infection, we studied the effects of antioxidant enzymes and hydroxyl radical scavengers on pulmonary clearance processes. Intratracheal injection of catalase and superoxide dismutase resulted in prolonged intraalveolar residence of the enzymes, but caused no decrease in rates of clearance of either Staphylococcus aureus 502A or Pseudomonas aeruginosa PAO1. In contrast, dimethylsulfoxide and dimethylthiourea caused significant depression of clearance of P. aeruginosa without altering clearance of S. aureus. These results provide further differentiation between clearance processes affecting gram-negative and gram-positive bacteria and suggest that murine clearance of gram-negative organisms may be in part mediated by reactions which generate hydroxyl anion. In vivo administration of agents which inhibit hydrogen peroxide-, superoxide-, or hydroxyl anion-mediated reactions do not alter normal clearance of S. aureus.
Structural and antigenic analysis of meningococcal piliation
Authors: Olafson R W , McCarthy P J , Bhatti A R , Dooley J S , Heckels J E , Trust T J
Published: January 2020

Pilin with an Mr of 16,500 was purified to homogeneity from Neisseria meningitidis SP3428. Procedures which provided useful separation during purification included high-pressure liquid chromatography with a TSK size exclusion column, Sephacryl S-200 column chromatography, ion-exchange chromatography with SP-Sephadex, and preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The amino acid composition of this pilin was similar to that previously reported for this species. The sequence of N-terminal 51 amino acids was also determined. The protein lacked a modified phenylalanine at the amino terminus and displayed six residues which were different from Neisseria gonorrhoeae in that region of the molecule determined to be the lectin-binding domain. Monoclonal antibody raised to this pilin was employed, along with a monoclonal antibody to an epitope common to all gonococcal pilins, to analyze the intra- and interstrain heterogeneity of meningococcal piliation. The results indicate that N. meningitidis displays considerable intra- and interstrain heterogeneity with respect to both pilus subunit size and antigenicity. The Mr of subunits ranged from 13,000 to 20,000.
Suppression of antigen- and mitogen-induced proliferation of bovine lymphocytes by excretory-secretory products of Oesophagostomum radiatum
Authors: Gasbarre L C , Romanowski R D , Douvres F W
Published: January 2020

Excretory-secretory products (ESP) isolated from in vitro-grown stage-3 to -4 larvae of Oesophagostomum radiatum were found to inhibit both the in vitro antigen-specific proliferation of keyhole limpet hemocyanin- and ovalbumin-primed lymphocytes and the proliferation induced by the T-cell mitogen concanavalin A. As little as 50 ng of ESP protein per culture resulted in 50% reductions of subsequent proliferative responses. Antigen-induced responses were 100 to 1,000 times more sensitive to inhibition than were mitogen-induced responses. The inhibitory activity was found to affect the induction of proliferation as evidenced by the observation that complete inhibition was seen when ESP were added to cultures within the first 24 h. ESP were found to have no inhibitory activity when added 72 h after the initiation of the cultures. The inhibition was not a result of a direct action upon macrophages because pulsing of adherent cells with ESP had no more effect on a subsequent proliferative response than did a pulsing of the culture vessel itself. The inhibitory activity eluted from high-pressure liquid chromatography columns in the same fractions as protein standards with molecular weights of 25,000 to 35,000. Of special interest is the fact that this inhibitor of the expansion of immunoreactive clones of lymphocytes is found associated with the stages of parasites most intimately associated with host tissues, namely larval stages 3 and 4.
Augmentation of interleukin-2 production and delayed hypersensitivity in mice infected with Mycobacterium bovis and fed a diet supplemented with vitamin A acetate
Authors: Colizzi V , Malkovsky M
Published: January 2020

High-dose Mycobacterium bovis-infected mice fed a vitamin A acetate-supplemented diet developed a positive skin reaction to purified protein derivative of mycobacteria, and their spleen cells showed an increased IL-2 production in vitro.
Effects of transmission-blocking monoclonal antibodies on different isolates of Plasmodium falciparum
Authors: Graves P M , Carter R , Burkot T R , Rener J , Kaushal D C , Williams J L
Published: January 2020

The strain diversity in Plasmodium falciparum has been studied with respect to gamete surface antigens which are the targets of transmission-blocking antibodies. Of 12 isolates tested, 11 were positive by immunofluorescence with the three monoclonal antibodies studied. The exception was a Liberian isolate, two clones of which were found to react with only one of the three monoclonal antibodies. Antibodies IIC5-B10 and IA3-B8, which previously have been shown to act synergistically to block infectivity of 7G8, a Brazilian clone of P. falciparum, acted in an exactly similar way with another Brazilian isolate, It.D12, and an isolate from Thailand. In the presence of complement either IA3-B8 or a third antibody, IID2-A10, strongly suppressed infectivity of It.D12 as well as 7G8, but neither isolate was strongly suppressed by IIC5-B10. IA3-B8 and IID2-A10 did not react by immunofluorescence or immunoprecipitation with gametes of L.E5; IIC5-B10 reacted positively with L.E5 gametes in these tests. In the absence of complement, the combination of IA3-B8 and IIC5-B10 did not suppress infectivity of L.E5 to mosquitoes. In contrast to its effect on gametocytes of other isolates, IIC5-B10 in the presence of complement strongly suppressed infectivity of L.E5 to mosquitoes. These results imply that IA3-B8 and IIC5-B10 react with two structurally distinct epitopes on the surface of gametes of P. falciparum and that the properties of both epitopes on gametes of L.E5 differ from those on gametes of the other isolates tested.
Immunological relationship of the B subunits of Campylobacter jejuni and Escherichia coli heat-labile enterotoxins
Authors: Klipstein F A , Engert R F
Published: January 2020

The application of dissociation techniques, involving gel filtration in the presence of guanidine, to a semipurified preparation of Campylobacter jejuni heat-labile enterotoxin yielded a material whose functional and immunological properties resemble those of the B subunits of cholera toxin and Escherichia coli heat-labile toxin (LT). The C. jejuni toxin B subunit reacted with GM1 ganglioside in an enzyme-linked immunosorbent assay, but lacked the holotoxin's cytotonic activity in the Chinese hamster ovary tissue culture assay and its ability to cause fluid secretion in rat ileal ligated loops. The C. jejuni toxin B subunit showed lines of partial identity with the B subunits of both cholera toxin and LT in gel immunodiffusion; it appeared to be more closely related immunologically to the LT B subunit than to the cholera toxin B subunit in enzyme-linked immunosorbent assays that used antisera either to LT or to its B subunit. Rats immunized with LT B subunit were significantly protected against challenge with either the semipurified C. jejuni toxin or a viable enterotoxigenic strain of C. jejuni, although twice the immunization dosage was required to achieve protection comparable to that against the homologous toxin or viable bacteria. These observations indicate that the C. jejuni enterotoxin contains a B subunit that bears an immunological relationship with the B subunits of cholera toxin and LT.
Bacterial lipopolysaccharide induction of leukocyte-derived corticotropin and endorphins
Authors: Harbour-McMenamin D , Smith E M , Blalock J E
Published: January 2020

Previous reports have shown that there is an endogenous opioid component associated with pathophysiological responses to endotoxin. It has been shown that these responses are alleviated by naloxone, a specific opiate antagonist. Results of another study have indicated that leukocytes may mediate some of those responses since leukocyte depletion alleviated the effects of lipopolysaccharide. In view of the above reports as well as the finding that leukocytes produce immunoreactive (ir-) endorphins and corticotropin (ACTH) when stimulated with Newcastle disease virus or ACTH-releasing factor, we postulated that leukocytes may serve as an extrapituitary source of endorphins produced in response to bacterial endotoxin. To test this hypothesis, human peripheral blood leukocytes as well as mouse spleen cells were cultured in vitro with Escherichia coli lipopolysaccharide for 48 h. The lipopolysaccharide (i.e., endotoxin) was shown to induce de novo synthesis of ir-ACTH and ir-endorphins. The leukocyte-derived ir-ACTH had a molecular weight of approximately 2,900 and demonstrated a bioactivity similar to that of pituitary-derived ACTH. The lymphocyte-derived ir-endorphin comigrated with alpha- and gamma-endorphin at approximately 1,800 daltons and was shown to bind to brain opiate receptors. These findings imply that leukocyte-derived endorphins may be involved in the pathophysiological response to endotoxin.
Biologically active antibodies elicited by a synthetic circumsporozoite peptide of Plasmodium knowlesi administered in saline with a muramyl dipeptide derivative
Authors: Clough E R , Audibert F M , Barnwell J W , Schlesinger D H , Arnon R , Chedid L A
Published: January 2020

A synthetic peptide whose sequence was derived from the circumsporozoite protein of Plasmodium knowlesi coupled to bovine gamma globulin has been shown to be immunogenic when administered with Freund complete adjuvant. The present experiments were designed to test the immunogenicity of the peptide when attached to a tetanus toxoid carrier and administered with alum or murabutide, both acceptable clinical adjuvants. In both cases, the use of an adjuvant increased the levels of circulating anti-peptide antibodies over those observed when no adjuvant was used. However, when the antisera were tested for reactivity with the native protein, animals of the group receiving the conjugate associated with murabutide always had titers greatly exceeding those observed in animals that received the conjugate with alum. Moreover, the sera of the murabutide-treated group were shown to be more active in eliciting shedding of the circumsporozoite protein than were sera of animals of the Freund complete adjuvant-treated group. The use of tetanus toxoid in secondary immunizations could be eliminated when the mice primed with peptide-tetanus toxoid and murabutide were boosted with a polymer of the peptide. The results indicate that the synthetic malarial peptide-tetanus toxoid conjugate is capable of stimulating high levels of biologically active antibodies only when administered with murabutide.
Reactivity of type-specific monoclonal antibodies with Staphylococcus aureus clinical isolates and purified capsular polysaccharide
Authors: Nelles M J , Niswander C A , Karakawa W W , Vann W F , Arbeit R D
Published: January 2020

Staphylococcus aureus has been classified into at least eight different capsular types by using polyclonal rabbit antisera specific for their associated capsular polysaccharides. We produced and characterized monoclonal antibodies reactive with two serologically distinct capsular types, types 5 and 8, which account for more than 70% of all S. aureus bacteremias. These type-specific, monoclonal antibodies reacted with S. aureus clinical isolates possessing the homologous capsular type and exhibited no cross-reactivity against S. aureus clinical isolates possessing the heterologous capsular type, nontypeable S. aureus clinical isolates, Staphylococcus epidermidis clinical isolates, or a variety of gram-negative organisms. The anti-type 8 monoclonal antibodies also reacted with purified capsular polysaccharide derived from the prototype type 8 S. aureus strain.
Cloning of plasmid DNA sequences involved in invasion of HeLa cells by Shigella flexneri
Authors: Maurelli A T , Baudry B , d'Hauteville H , Hale T L , Sansonetti P J
Published: January 2020

A large plasmid is found in virulent isolates of Shigella sp. and encodes functions essential for invasion of mammalian cells. To identify plasmid sequences necessary for invasion, we isolated a series of Tn5 insertions in pWR100, the virulence plasmid of Shigella flexneri serotype 5. These insertions demonstrated that three separate EcoRI fragments of pWR100 were required for invasion of HeLa cells. However, the corresponding native EcoRI fragments, when cloned into pBR325, did not restore virulence to plasmidless strains. Construction of a lambda-sensitive, plasmidless Shigella recipient enabled us to shotgun clone plasmid DNA directly into S. flexneri by using the cosmid vector pJB8 and score for expression of invasive functions. In this fashion, we succeeded in isolating six independent recombinants which restored invasion of HeLa cells in plasmidless Shigella recipients. The cloned inserts all contained a common core of ca. 37 kilobases, thus defining a minimum sequence necessary for invasion of HeLa cells. Virulence-associated peptides produced by wild-type S. flexneri were also produced by the recombinants. Expression of these peptides and expression of invasiveness by the clones were regulated by growth temperature, as is expression of these traits in wild-type S. flexneri. A complete invasive phenotype was not expressed by the recombinants in that they failed to produce a positive Sereny test. Possible explanations for this behavior as it relates to the mechanism of bacterial invasion are discussed.
Effect of estrogen (17 beta-estradiol) on the susceptibility of mice to disseminated gonococcal infection
Authors: Kita E , Takahashi S , Yasui K , Kashiba S
Published: January 2020

Studies of the effect of sex hormones on the susceptibility of mice to the disseminated gonococcal infection demonstrated significantly enhanced susceptibility of mice injected with estrogen (17 beta-estradiol). In mice treated with estradiol, bacteremia progressively developed within 12 h postinoculation and mice died within the next 6 h, whereas bacteremia in mice treated with progesterone was completely cleared within 3 h postinoculation. The administration of estradiol affected the function of polymorphonuclear leukocytes (PMN) responsible for eliminating gonococci, but the administration of progesterone did not. The bactericidal activity of PMN mediated by myeloperoxidase was affected by estradiol, but the capacity of PMN to release superoxide anion was not. Furthermore, peritoneal cell analysis demonstrated that the infiltration of PMN in the peritoneal cavity of estradiol-treated mice significantly decreased when mice were injected intraperitoneally with gonococci. These effects on PMN by estradiol may play an important role in the enhanced susceptibility of estradiol-treated mice to gonococcal infection.
Immune suppression and induction of gamma interferon by pertussis toxin
Authors: Vogel F R , Klein T W , Stewart W E , Igarashi T , Friedman H
Published: January 2020

It has been suggested that pertussis toxin is a virulence factor of Bordetella pertussis. Although extracts enriched in pertussis toxin activity have been reported to enhance immune responsiveness, other studies have demonstrated a suppressive ability, suggesting that the toxin may contribute to the virulence of B. pertussis through mechanisms involving immune suppression. We report that purified pertussis toxin suppressed the in vitro immunoglobulin M antibody response of mouse splenocytes to sheep erythrocytes. At submitogenic doses, the toxin also suppressed [3H]thymidine incorporation by splenocytes, suggesting that it interfered with antibody formation by inhibiting lymphocyte proliferation. Antiviral activity was detected in culture supernatants obtained from pertussis toxin-suppressed splenocyte cultures by using a cytopathic effect inhibition assay. This antiviral activity was virus nonspecific, sensitive to pH 2.0 treatment, stable to heating at 56 degrees C, and neutralized by anti-gamma interferon antiserum. Finally, the fractionation of splenocytes by anti-immunoglobulin panning techniques suggested that Lyt2+ lymphocytes proliferated in response to pertussis toxin and produced interferon. Our results suggest that pertussis toxin may contribute to the virulence of B. pertussis through stimulation of Lyt2+ lymphocytes, resulting in the induction of gamma interferon and the subsequent inhibition of the primary antibody response.
Activation and inhibition of Limulus amebocyte lysate coagulation by chemically defined substructures of lipid A
Authors: Proctor R A , Textor J A
Published: January 2020

Recent work with lipid mutants of Escherichia coli and Salmonella typhimurium has helped to elucidate the correct structure of lipid A and has suggested a biosynthetic pathway. Precursor molecules include diacylglucosamine 1-phosphates and tetraacyl disaccharide bis-phosphates. The activities of several of these compounds and of their derivatives were measured by Limulus amebocyte lysate (LAL) assay. We report that (i) both mono- and disaccharide precursors of lipid A activate LAL, (ii) two acyl chains on the monosaccharide subunit of lipid A are necessary for activation of LAL, and (iii) the monosaccharide, 2-monoacylglucosamine 1-phosphate can competitively inhibit LAL activation by diacyl monosaccharide lipid A precursors. However, 2-monoacylglucosamine 1-phosphate did not inhibit endotoxin activation of LAL. One unanticipated finding was that the activities of the monosaccharides were reduced upon storage even though their covalent structures were unchanged. Perhaps this is due to alterations in physical state. Thus, these lipid A precursors and derivatives offer some insight into the structural features required for activation of the LAL assay and may in the future provide derivatives which are competitive inhibitors of endotoxin.
Genetics of spontaneous, high-frequency loss of b capsule expression in Haemophilus influenzae
Authors: Hoiseth S K , Connelly C J , Moxon E R
Published: January 2020

We determined that the frequency of spontaneous capsule loss in Haemophilus influenzae type b is 0.1 to 0.3%. All of 10 independent capsule-deficient variants (derived from four different type b strains) were found to be missing an identical 9-kilobase EcoRI restriction fragment when probed with a cloned piece of DNA containing sequences known to be necessary for type b capsule expression. These results suggest the existence of a specific mechanism for shutting off type b capsule synthesis at a high frequency. Intranasal infection of infant rats showed that capsule loss occurred in vivo at frequencies comparable to those observed in vitro.
Specific suppressor T cells for delayed-type hypersensitivity in susceptible mice immunized against cutaneous leishmaniasis
Authors: Dhaliwal J S , Liew F Y , Cox F E
Published: January 2020

BALB/c mice injected intravenously with 10(6) or higher doses of formaldehyde-fixed promastigotes (ffp) of Leishmania major developed significantly lower levels of delayed-type hypersensitivity (DTH) compared with uninjected control mice when they were subsequently immunized intradermally with ffp. The suppression of DTH was antigen specific and was also inducible with lethally irradiated promastigotes or soluble parasite antigens. The suppressive effect was adoptively transferable with splenic T cells which express the Lyt-1+2+ and L3T4+ phenotypes. These specific suppressor T cells were active against both the inductive and expressive phases of DTH. They were sensitive to 200 rads of gamma-irradiation in vitro and appeared to manifest the suppressive activity via soluble factors. In spite of this profound suppression of DTH, BALB/c mice injected intravenously with 4 X 10(7) ffp were substantially protected against a challenge infection with L. major promastigotes. The possible relationship between the suppressor T cells for DTH and prophylactic immunization against fatal cutaneous leishmanial infection in susceptible BALB/c mice is discussed.
Induction of protective immunity to Plasmodium falciparum in Saimiri sciureus monkeys with partially purified exoantigens
Authors: James M A , Kakoma I , Ristic M , Cagnard M
Published: January 2020

Soluble Plasmodium falciparum exoantigens in crude culture supernatant fluids induced protective immunity against experimental falciparum malaria in Bolivian Saimiri sciureus monkeys. Susceptible squirrel monkeys were vaccinated with an aluminum hydroxide-fortified fraction purified from culture supernatants of P. falciparum Indochina I and Geneve/SGE-1 by cation-exchange (sulfopropyl-trisacryl) chromatography. Animals immunized with sulfopropyl-purified and corresponding control immunogens were challenged with whole blood containing monkey-adapted virulent organisms of the Indochina I strain. Hematological, serological, and parasitological profiles, including the appearance of crisis forms, served as potential indicators of protection. This immunogen conferred significant clinical protection of squirrel monkeys against needle challenge with the homologous Indochina I strain and a moderate degree of heterologous strain immunity.
Protection of rabbits against experimental pasteurellosis by vaccination with a potassium thiocyanate extract of Pasteurella multocida
Authors: Ringler D H , Peter G K , Chrisp C E , Keren D F
Published: January 2020

Antigens were extracted from a virulent isolate of Pasteurella multocida (serotype 3, 12, 15:D) with potassium thiocyanate, and a vaccine was prepared. Pasteurella-free rabbits were vaccinated intranasally and intraconjuctivally twice with a 2-week interval and challenged intranasally with the homologous P. multocida serotype 2 weeks after the second vaccination. The vaccinated rabbits produced serum immunoglobulin G and nasal mucosal immunoglobulin A against P. multocida. The vaccine protected the challenged rabbits against clinical disease and death; however, otitis media was not prevented, and microscopic inflammatory lesions were occasionally noted in the lungs and nasal turbinates. In contrast, nonvaccinated, challenged rabbits became febrile, dyspnic, depressed, and anorectic, and five of six died within 4 days of challenge with severe lesions including pneumonia, pleuritis, otitis media, and bacteremia. The vaccine prevented death and colonization of challenge organisms in the blood and lung, but did not prevent colonization of the middle ear. The vaccine alone did not cause clinical disease or gross lesions, but did produce microscopic pulmonary inflammatory lesions.
Genetic evidence for role of extracellular proteinase in virulence of Candida albicans
Authors: Kwon-Chung K J , Lehman D , Good C , Magee P T
Published: January 2020

The relationship between extracellular proteinase and the virulence for mice in Candida albicans was studied by using a set of three isolates. The set included a proteinase-producing parent (C9), a proteinase-deficient mutant derived from C9 by nitrous acid treatment (C9M1), and a spontaneous revertant (C9M1M) obtained by mouse passage of C9M1. The morphological markers and the carbon assimilation pattern were identical in these isolates. Isolate C9 produced a high level of proteinase in vitro and caused fatal infection (100%) within 21 days. The mutant produced no detectable enzymes in vitro, and all mice survived until day 22. Only 30% of the mice infected with C9M1 died between day 23 and 30. The isolates recovered from the dead mice were found to be proteinase sufficient, indicating that the mice died after the organism in tissue had reverted. The C9M1M isolate produced proteinase in vitro at 44% the level of C9 and induced fatal infection in 90% of the mice within 30 days. The number of CFU recovered from the kidneys correlated with the level of proteinase produced in vitro and, in turn, the rate of fatal infection produced by the isolates. These results support a previous observation indicating that proteinase activity is one of the virulence factors associated with C. albicans.
Immunochemical characterization of P pili from invasive Escherichia coli
Authors: Hanley J , Salit I E , Hofmann T
Published: January 2020

P pili (or fimbriae) are present on most pyelonephritogenic Escherichia coli strains, and they mediate binding to erythrocytes and epithelial cells. To determine the antigenic diversity of P pili, we purified the pili from 14 bacteremic E. coli strains which caused mannose-resistant hemagglutination. Pilus preparations consisted of one to three bands in sodium dodecyl sulfate-polyacrylamide gel electrophoresis and ranged in molecular weight from 14,000 to 19,500. There was no single band common to all the strains. An enzyme-linked immunosorbent inhibition assay detected 20 ng of pilus antigen. When four different rabbit antisera were used, only two or fewer heterologous strains could inhibit the enzyme-linked immunosorbent assay. Immunoblots yielded the same results. Protein sequences of four P pili had identical N termini. These results show that despite having identical amino-terminal sequences, P pili are antigenically heterogeneous. The receptor-binding domains which are likely to be identical in all strains must be immunorecessive.
Analysis of effects of lipopolysaccharide and interferon on murine macrophages: modulation of elastase secretion in vitro
Authors: Duc Dodon M , Vogel S N
Published: January 2020

Macrophages are able to produce and secrete elastase in response to a variety of agents which induce macrophage differentiation. In this study, we compared elastase levels in macrophage cultures derived from lipopolysaccharide (LPS)-responsive and LPS-hyporesponsive mice. After the in vivo administration of fluid thioglycolate, both types of macrophages exhibited increased secretion of elastase, and further enhancement was observed after in vitro stimulation with colchicine or phorbol myristic acetate or ingestion of latex beads. In contrast, phenol- and water-extracted, protein-free preparations of LPS (Ph-LPS) markedly inhibited elastase secretion below basal levels in LPS-responsive macrophages. The LPS-induced inhibition was reversible with polymyxin B and was not observed in Ph-LPS-stimulated C3H/HeJ (LPS-hyporesponsive) macrophage cultures. Stimulation of either LPS-responsive or -hyporesponsive macrophage cultures with interferon (IFN) also resulted in a significant reduction in elastase secretion below basal levels. LPS-induced inhibition of elastase secretion could be reversed and elastase secretion could be augmented in the presence of an antibody directed against IFN-alpha/beta. These findings suggest that LPS induces the production of both elastase and IFN, and that the latter product acts to suppress secretion of the proteinase.
Electrophoretic separation and molecular weight characterization of Pseudomonas aeruginosa H-antigen flagellins
Authors: Allison J S , Dawson M , Drake D , Montie T C
Published: January 2020

We found that preparations of Pseudomonas aeruginosa flagellar antigens protected against P. aeruginosa challenge in a burned-mouse model. To determine the extent of similarity among known flagellar antigen types, we compared flagellins by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. A majority of our laboratory strains and clinical isolates, including PAO strains and their derivative mutants RM46 and PJ108, as well as virulent strains M-2 and 1244, had flagellins of 53,000 Mr. These flagellins had the same Mrs as those of type b-standard strains 170001 and 15084. The heterogeneous group of a-type H-antigen flagellins were of smaller molecular weights, ranging from 52,000 Mr for standard strain 5939 (a0, a3) to 45,000 Mr for standard strain 170018 (a0, a3, a4). Standard strains 5933 (a0, a1, a2) and 5940 (a0, a2) had intermediate Mrs of 51,000 and 47,000, respectively. Differences in Mr of 1,000 to 2,000 could be resolved by coelectrophoresis. A series of 26 unknown strains were categorized. Correlations among typing by molecular weight, cross-agglutination reactions with O-adsorbed H antisera, and previous results for H-serum typing are reported.
Putative Treponema pallidum cytadhesins share a common functional domain
Authors: Thomas D D , Baseman J B , Alderete J F
Published: January 2020

Three putative Treponema pallidum ligands (P1, P2, and P3) that bind host fibronectin were characterized by peptide mapping. Papain digestion of each protein yielded a comigrating peptide of approximately 12,000 molecular weight. An antibody to this protein fragment inhibited T. pallidum host cytadherence, indicating that this peptide may be the functional domain of these treponemal adhesins.
Factors Affecting Virulence of
Shigella flexneri:
Strain with Relaxed Control of Ribonucleic Acid Synthesis
Authors: Rothman Sara W. , Corwin Laurence M.
Published: January 2020

An
Escherichia coli-Shigella flexneri
hybrid strain which produces an abortive, nonfatal experimental dysentery has inherited relaxed control of ribonucleic acid synthesis from its
E. coli
parent. This property, coupled with a partial methionine requirement, has been used to explain its long lag phase in vitro, relative to its
Shigella
parent. It was suggested that such characteristics may contribute to the hybrid's poor survival in the gut resulting in the abortive infection.
Comparison of Capsular Types of
Staphylococcus aureus
Strains
Authors: Yoshida Kosaku , Naito Yoshiko
Published: January 2020

Staphylococcal capsular types 1 to 4 as determined by the “specific capsular reaction” belonged to type A capsule as determined by the serum-soft agar technique.
Chemical Properties of the Pili of
Corynebacterium renale
Authors: Kumazawa Norichika , Yanagawa Ryo
Published: January 2020

Pili separated from the cells of
Corynebacterium renale
strain 46 (type II) by agitation at high speed in a homogenizer were purified by repeated cycles of ammonium sulfate precipitation, sonic treatment, and centrifugation. The preparation of purified pili formed a single antigen-antibody line in agar gel and showed an absorption maximum at 275 nm. The pili subjected to dodecyl sulfate-polyacrylamide gel electrophoresis formed a main band and corresponded to the molecular weight of 19,000. The fact that the total nitrogen of the amino acids of the pili was nearly equal to its nitrogen content, together with the absence of detectable carbohydrate, has led to the conclusion that the pili are protein. The pilial protein was composed of 20 amino acids. Preparations of pili which had been treated with 0.5
n
NaOH, but not with 1
n
HCl, no longer appeared filamentous and failed to form a precipitate with the antibody in agar gels. A comparison has been made of the amino acid composition and certain properties of the pili of
C. renale
and type I pili and F pili of gram-negative bacteria.
Prolonged Stationary-Phase Growth of T-Strain Mycoplasmas in Broth Culture
Authors: Hendley J. Owen , Allred Elizabeth N.
Published: January 2020

Growth of T-strain mycoplasmas in standard broth cultures has been characterized by rapid growth to peak titers of 10
6
to 10
7
color change units/ml at 20 to 24 hr, followed by a sharp decline in the viability over the next few hours. In a vacuum-flow (V-F) system utilizing negative pressure equal to 10 cm of water and air flow on the surface of broth, T strains were seen to enter the stationary phase of growth after 24 hr, and viability titers after 72 hr were ≥ 10
4
color change units/ml. Maintenance of organisms in the stationary phase required both vacuum and air flow, a medium depth of 1 mm, and 1% urea in the broth. Concentration of ammonia in broth cultures under V-F remained below 200 μg/ml during 72 hr of observation. Ammonia levels in standard broth cultures exceeded 200 μg/ml after 24 hr, coincident with the decline in viability. Air passed over the medium surface in the V-F system contained large amounts of ammonia; the amount increased during each succeeding 24-hr period of observation.
Use of Specifically Labeled Sucrose for Comparison of Extracellular Glucan and Fructan Metabolism by Oral Streptococci
Authors: Schachtele C. F. , Loken A. E. , Schmitt M. K.
Published: January 2020

Sucrose labeled in the fructosyl (
3
H) and glucosyl (
14
C) moieties was used to quantitate extracellular polysaccharide production and degradation by cariogenic and noncariogenic oral streptococci. All of the strains produced glucan and fructan.
Streptococcus salivarius
produced primarily fructan, whereas
S. mutans
and
S. sanguis
produced more glucan than fructan. The cariogenic streptococci could degrade the fructan produced by noncariogenic strains. Although the soluble glucans from all of the strains were sensitive to dextranase, the insoluble glucan from
S. mutans
could be distinguished from the
S. sanguis
insoluble glucan by its greater resistance to this enzyme.