Identification of Proteus penneri sp. nov., formerly known as Proteus vulgaris indole negative or as Proteus vulgaris biogroup 1.

The name Proteus penneri sp. nov. is proposed for a group of organisms previously called Proteus vulgaris indole negative or P. vulgaris biogroup 1. All of these strains were salicin negative, esculin negative, and chloramphenicol resistant (zone size, less than 14 mm). DNA relatedness studies indicated that when DNA from P. penneri strain 1808-73 was labeled and tested against unlabeled DNA from 13 other P penneri strains, a highly related group was formed (88 to 99% relatedness at 60 degrees C and 67 to 99% relatedness at 75 degrees C). Strain 1808-73 (ATCC 33519) is proposed as the type strain of P. penneri. In this study, two distinct groups of indole-positive P. vulgaris strains were also apparent. The first group (defined as P. vulgaris biogroup 2) was indole positive, salicin positive, and esculin positive, and the second group (defined as P. vulgaris biogroup 3) was indole positive, salicin negative, and esculin negative. The current type strain of P. vulgaris (ATCC 13315) belongs to biogroup 3. The DNA from P. penneri strains was not highly related to labeled DNA from the type strain of P. vulgaris (14 to 30% relatedness at 75 degrees C) or from P. vulgaris strain PR 1 (ATCC 29905), which belongs to biogroup 2 (27 to 33% relatedness at 75 degrees C). Strains of biogroup 2 were sensitive to chloramphenicol (zone size, greater than 19mm), and 10 of these strains formed a highly related group by DNA hybridization when DNA from PR 1 was labeled (64 to 100% relatedness at 60 degrees C and 70 to 100% relatedness at 75 degrees C), but they were not highly relatedness to the type strain of P. vulgaris (51 to 68% relatedness at 60 degrees C and 14 to 44% relatedness at 75 degrees C). Further DNA relatedness studies are needed on strains of biogroup 3 before a definitive taxonomic proposal can be made for these two indole-positive biogroups.     

Formation of rabbit reaginic antibodies to protein and hapten—protein conjugates

The capacities of BSA and DNP—protein conjugates to evoke reagin formation in rabbits were compared. Reagins to DNP generally appeared earlier and disappeared more rapidly from the circulation than did anti-BSA reagins. Initial formation of reagins proceeded with a logarithmic phase indicating a doubling time of 7–8 hours. Booster antigen injections resulted in some cases in a reagin response after a shorter latent phase than that observed after primary immunization. A secondary reagin response was more readily evoked in rabbits with low titres of agglutinating antibodies than in those with high titres. Anti-DNP reagins were demonstrable in a higher percentage of the injected rabbits than were anti-BSA reagins. The two types of reagins were equally sensitive to heat and 2-mercaptoethanol. A positive correlation between serum levels of anti-DNP but not anti-BSA reagins and agglutinating antibodies was demonstrated. Some evidence that a low antigen dose was more efficient than a high dose in evoking reagin formation was obtained. Treatment of rabbits with 6-mercaptopurine during the 1st week following antigen injection resulted in an increased latent phase and an enhancement of the production of anti-BSA reagins and some suppression of the formation of anti-DNP reagins.     

The effect of sympathetic stimulation on proximal brachial artery mechanics in humans--differential behaviour within the length of the brachial artery?

AIMS: The mechanical properties of arteries play a major role in the regulation of blood pressure and cardiac performance. The effect of sympathetic stimulation on the mechanical properties of the proximal brachial artery was analysed in 18 healthy volunteers, nine young (25 +/- 2 years) and nine elderly (69 +/- 2 years). METHODS: A non-invasive ultrasonic echo-tracking system for measurement of systolic/diastolic variation of the proximal brachial artery diameter in combination with intra-arterial pressure measurements was used to determine wall mechanics. The pressure-diameter (P-D) relationship, distensibility coefficient (DC), compliance coefficient (CC) and stiffness(beta) were obtained at rest and during sympathetic stimulation induced by lower body negative pressure (LBNP). RESULTS: The peripheral vascular resistance increased by 100 and 72%, respectively in the young and elderly during LBNP (P < 0.001). Simultaneously, the mechanical properties of the proximal brachial artery remained unaltered, as estimated from both P-D relationship and stiffness in young (beta-index rest: 5.2 +/- 0.9, LBNP: 5.5 +/- 1.3, NS) as well as elderly (beta-index rest: 13.6 +/- 4.6, LBNP: 16.1 +/- 4.7, NS). CONCLUSIONS: LBNP-induced sympathetic activation does not change proximal brachial artery mechanics, in contrast to earlier reports on the muscular distal brachial artery. This may imply that the transition between elastic and muscular artery behaviour is within the length of the brachial artery, where the site of transition from elastic to muscular wall structure needs to be specified in future studies.     

山莨菪碱对家兔动脉粥样硬化的抑制作用

用65只日本兔喂高脂饲料,观察山莨菪碱对动脉粥样硬化形成的作用及其有关检测指标的影响。结果表明,山莨菪碱组主动脉内膜粥样硬化斑块面积、胆固醇含量、内膜通透性及血液TC、LDL-C、TG、LPO、TXA_2、5-HT、Pt聚集力,血液流变学等,均显著降低;PGI_2及SOD增高;组织光镜及电镜改变减轻。说明山茛菪碱对动脉粥样硬化的形成有显著抑制作用。     

In Vitro and In Vivo Interactions of Haemophilus ducreyi with Host Phagocytes

We investigated the phagocytosis of Haemophilus ducreyi both in vitro and in vivo. Human granulocyte and monocyte phagocytosis of opsonized and nonopsonized, fluorescence-labeled H. ducreyi was assessed by flow cytometry. Both Escherichia coli and noncapsulated H. influenzae were included as controls. The maximal percentage of granulocytes taken up by H. ducreyi was 35% after 90 min. In contrast, 95% of H. influenzae bacteria were phagocytosed by granulocytes after 30 min. These results indicated that H. ducreyi phagocytosis was slow and inefficient. Bacterial opsonization by using specific antibodies increased the percentage of granulocytes phagocytosing H. ducreyi from 24 to 49%. The nonphagocytosed bacteria were completely resistant to phagocytosis even when reexposed to granulocytes, indicating that the H. ducreyi culture comprised a mixture of phenotypes. The intracellular survival of H. ducreyi in granulocytes, in monocytes/macrophages, and in a monocyte cell line (THP-1) was quantified after application of gentamicin treatment to kill extracellular bacteria. H. ducreyi survival within phagocytes was poor; approximately 11 and <0.1% of the added bacteria survived intracellularly after 2 and 20 h of incubation, respectively, while no intracellular H. influenzae bacteria were recovered after 2 h of incubation with phagocytes. The role of phagocytes in the development of skin lesions due to H. ducreyi was also studied in vivo. Mice that were depleted of granulocytes and/or monocytes and SCID mice, which lacked T and B cells, were injected intradermally with approximately 10⁶ CFU of H. ducreyi. Within 4 days of inoculation, the granulocyte-depleted mice developed lesions that persisted throughout the experimental period. This result reinforces the importance of granulocytes in the early innate defense against H. ducreyi infection. In conclusion, H. ducreyi is insufficiently phagocytosed to achieve complete eradication of the bacteria. Indeed, H. ducreyi has the ability to survive intracellularly for short periods within phagocytic cells in vitro. Since granulocytes play a major role in the innate defense against H. ducreyi infection in vivo, bacterial resistance to phagocytosis probably plays a crucial role in the pathogenesis of chancroid.     

Characterization of murine monoclonal antibodies to Escherichia coli J5.

Results show that various inbred strains of mice can be segregated into two distinct groups, based on their capacity to allow a number of nontuberculous mycobacterial infections to grow in target organs following experimental intravenous infection. The first group, which allowed these infections to grow progressively, was thus designated as naturally susceptible to these infections; in contrast, those strains which were able to exert detectable bacteriostasis were designated as naturally resistant. It was then found that segregation of mouse strains based on this distinction also mirrored the capacity of these animals to generate acquired immunity to the mycobacterial infections. For example, Mycobacterium simiae grew progressively in susceptible C57BL/6 mice, subsequently triggering acquired mechanisms of immunity, whereas no evidence for acquired immunity could be found in resistant A/Tru mice infected with this organism. The possibility that acquired immunity could not be expressed in the latter strain as a result of a defect in macrophage activation was excluded. Moreover, it was found that the trait of resistance to these infections could be transferred by bone marrow cells into radiation chimeras, thus indicating that this trait was expressed by the progeny of hemopoietic precursor cells. Subsequent backcross analysis to determine the mode of inheritance of the trait of resistance to these mycobacterial infections revealed data that were consistent with the hypothesis that this resistance is controlled by more than one gene. Statistical analysis of the data by the maximum likelihood method suggested polygenic control, although in some cases the probability values suggested control by a major gene, influenced by modifier genes. These findings suggest that the previous hypothesis that the growth of mycobacterial infections in inbred strains of mice is controlled by a single gene should be reevaluated.     

Bacteriophage Mu d1(Apr lac) generates vir-lac operon fusions in Shigella flexneri 2a.

Previous studies have demonstrated that expression of virulence in Shigella spp. is controlled by growth temperature. To study the regulation of virulence (vir) genes, we set out to develop a rapid, easily-assayed phenotype with which to measure expression of virulence. This report described a procedure for isolating vir-lac operon fusions in S. flexneri 2a by using the specialized transducing bacteriophage Mu d1(Apr lac) of Casadaban and Cohen (M. Casadaban and S. N. Cohen, Proc. Natl. Acad. Sci. U.S.A. 76:4530-4533, 1976). Mu d1(Apr lac) lysogens were isolated and screened for loss of virulence and for temperature-dependent expression of the lactose genes on Mu d1(Apr lac). A recombinant plasmid carrying the Mu immunity gene was also introduced into lysogens of interest to stabilize the Mu d1(Apr lac) insertion and prevent possible thermal induction at 37 degrees C. The mutant which we isolated failed to penetrate tissue culture cells in the assay for virulence and produced almost 15-fold more beta-galactosidase when grown at 37 degrees C than when grown at 30 degrees C. The site of insertion of Mu d1(Apr lac) in this strain was shown to be in the 140-megadalton plasmid pSf2a140, which is known to be associated with virulence. P1L4-mediated transduction of the insertion into a virulent recipient demonstrated genetic linkage of Mu d1(Apr lac) with loss of virulence and temperature-dependent expression of beta-galactosidase. All of these features fulfill the phenotype expected for a Mu d1(Apr lac)-induced vir-lac operon fusion. This mutant provides us with a means of measuring expression of a gene function required for virulence by assaying for beta-galactosidase. The insertion will also serve as a starting point for mapping of genes on pSf2a140 which are necessary for expression of virulence.     

Human immunoglobulin G antibody response to the major gonococcal iron-regulated protein.

In humans, gonococcal infection occurs in environments limited with respect to free iron. Neisseria gonorrhoeae produces increased quantities of iron-regulated membrane proteins when grown under in vitro conditions which restrict the availability of free iron. Using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot (immunoblot) techniques, we studied the reactivity of specific antibodies to the 37-kilodalton (kDa) major iron-regulated protein (MIRP) of gonococci grown under iron-limiting conditions. Antibodies reactive with the 37-kDa MIRP were distinguished from those reactive with protein I by using purified 37-kDa MIRP or gonococcal protein preparations. Acute-phase sera from patients with disseminated gonococcal infection (DGI) reacted strongly to both the 37-kDa MIRP and protein I. Acute sera from nine patients with uncomplicated gonorrhea did not exhibit strong reactivity with the 37-kDa MIRP and were indistinguishable from five control sera. When compared with acute-phase sera, convalescent-phase sera from patients with DGI failed to demonstrate increased reactivity, whereas convalescent-phase sera from one of nine patients with uncomplicated gonorrhea developed reactivity to the 37-kDa MIRP. These data indicate that (i) the 37-kDa MIRP is expressed and antigenic in vivo and (ii) humans with DGI consistently develop a systemic antibody response to the 37-kDa MIRP.     

Enterobacter asburiae sp. nov., a new species found in clinical specimens, and reassignment of Erwinia dissolvens and Erwinia nimipressuralis to the genus Enterobacter as Enterobacter dissolvens comb. nov. and Enterobacter nimipressuralis comb. nov.

Enterobacter asburiae sp. nov. is a new species that was formerly referred to as Enteric Group 17 and that consists of 71 strains, 70 of which were isolated from humans. Enterobacter asburiae sp. nov. strains gave positive reactions in tests for methyl red, citrate utilization (Simmons and Christensen's), urea hydrolysis, L-ornithine decarboxylase, growth in KCN, acid and gas production from D-glucose, and acid production from L-arabinose, cellobiose, glycerol (negative in 1 to 2 days, positive in 3 to 7 days), lactose, D-mannitol, alpha-methyl-D-glucoside, salicin, D-sorbitol, sucrose, trehalose, and D-xylose. They gave negative reactions in the Voges-Proskauer test and in tests for indole, H2S production, phenylalanine, L-lysine decarboxylase, motility, gelatin, utilization of malonate, lipase, DNase, tyrosine clearing, acid production from adonitol, D-arabitol, dulcitol, erythritol, i(myo)-inositol, melibiose, and L-rhamnose. They gave variable reactions in tests for L-arginine dihydrolase (25% positive after 2 days) and acid production from raffinose (69% positive after 2 days). Thirty-four Enterobacter asburiae sp. nov. strains were tested for DNA relatedness by the hydroxyapatite method with 32PO4-labeled DNA from the designated type strain (1497-78, ATCC 35953). The strains were 69 to 100% related in 60 degrees C reactions and 63 to 100% related in 75 degrees C reactions. Divergence within related sequences was 0 to 2.5%. Relatedness of Enterobacter asburiae sp. nov. to 84 strains of members of the Enterobacteriaceae was 5 to 63%, with closest relatedness to strains of Enterobacter cloacae, Erwinia dissolvens, Enterobacter taylorae, Enterobacter agglomerans, Erwinia nimipressuralis, and Enterobacter gergoviae. All strains tested were susceptible to gentamicin and sulfdiazine, and most were susceptible to chloramphenicol, colistin, kanamycin, nalidixic acid, carbenicillin and streptomycin. All strains were resistant to ampicillan, cephalothin, and penicillin, and most were resistant or moderately resistant to tetracycline. Enterobacter asburiae sp. nov strains were isolated from a variety of human sources, most prevalent of which were urine (16 strains), respiratory sources (15 strains), stools (12 strains), wounds (11 strains), and blood (7 strains). The clinical significance of Enterobacter aburiae is not known. As a result of this and previous studies, proposals are made to transfer Erwinia dissolvens and Erwinia nimipressuralis to the genus Enterobacter as Enterobacter dissolvens comb. nov. and Enterobacter nimipressuralis comb. nov., respectively.     

Epidemiological studies of Streptococcus pneumoniae in infants: methods of isolating pneumococci.

A prospective study of the natural history of pneumococcal infection, which involves serial culture studies in healthy infants from 6 weeks of age onward, is in progress in our laboratory. This report describes results of a comparison of several methods for the isolation and identification of Streptococcus pneumoniae from the nasopharynges and throats of these infants. Sheep blood agar, sheep blood agar with gentamicin sulfate (gentamicin agar), and mouse inoculation with 4-h broth cultures were used. Gentamicin agar proved superior to plain sheep blood agar as a solid culture medium, especially in enhancing the recovery of pneumococci from throat cultures. With gentamicin agar, similar carrier rates were found for both culture sites (nasopharynx and throat). In addition, gentamicin agar proved superior to mouse inoculation for the recovery of carrier strains from 131 nasopharyngeal culture samples processed by both methods. Sixty of 131 samples were positive for pneumococci, 25% of which would have been missed had mouse inoculation alone been used. In only three instances did we recover a strain by mouse inoculation that failed to grow on gentamicin agar; conversely, 15 strains were isolated on gentamicin agar but could not be recovered from mice. The latter observation might be explained by the fact that certain carrier strains may be relatively mouse avirulent. The use of blood agar containing gentamicin appears to offer a simple and inexpensive method for the recovery of S. pneumoniae and, in our opinion, provides an ideal method for the identification of pneumococcal carriers as well as for the recovery of these strains from clinical material such as sputum or ear exudates, where other and less fastidious organisms may also be present.