Resistance of respiratory and enteral bacteria to antibiotics

The relationship of the serum antibody titer and avidity to the putative periodontal pathogens Actinobacillus actinomycetemcomitans (Aa) strains Y4 and 29523 and Porphyromonas gingivalis (Pg) strain 381 were examined in relation to clinical parameters in 26 gingivitis and 28 periodontitis patients. The relationship of antibody titer and avidity to infection with the homologous organism was also examined in a subset of 30 patients. Antibody titer was determined by an enzyme-linked immunosorbent assay, and antibody avidity was assessed using a dissociation assay. Considering all patients, there was a significant negative correlation between mean probing depth and antibody titer (r=-0.28) and avidity (r=-0.28) to Aa Y4. There was a significant positive correlation of probing depth and antibody titer (r=0.46) and avidity (r=0.46) to Pg. The correlation of antibody titer and avidity to Aa and infection with Aa Y4 (r=-0.32, r=-0.21) and Aa 29523 (r=-0.35, r=-0.39) was negative, while the correlations of titer and avidity to Pg and presence of the organisms was strongly positive (r=0.40, r=0.35). These data indicate that the relationship of serum antibody titer and avidity to clinical parameters of periodontal disease severity and the level of infection with the homologous organism appears to be different for Aa and Pg. The development of an antibody response to Aa appears to protect the individual from infection with the organism. In contrast, the development of an antibody response to Pg was not able to eliminate the infection. These results should be considered when developing a diagnostic strategy for periodontal disease utilizing the humoral immune response.     

Resistance to antibiotics of Streptococcus pneumoniae strains

Streptococcus pneumoniae strains are exhibiting increasing rates of antibiotics resistance. A rapid increase of resistance was seen not only to penicillin but also other antimicrobial agents and therefore this paper describes the study of resistance and multiresistance of pneumococci to 7 antibiotics: penicillin (P), erythromycin (E), clindamycin (CC), tetracycline (T), co-trimoxazole (SXT), cefotaxime (CTX) and vancomycin (Va), using the disk-diffusion technique according to NCCLS procedure. We tested a total of 218 S. pneumoniae strains isolated from various materials: from sputum (54), noses (117), throats (28) and different swabs specimens (19). The overall percentage of resistant isolates to penicillin was 3.7%, to erythromycin--4.1%, to clindamycin--10.6%, to tetracycline--17.4%, to co-trimoxazole--15.6%, to cefotaxime--2.3%. In the sputum was most the monoresistant strains (66.7%). The multiresistance was highest in the penicillin resistant pneumococci. With the exception of vancomycin, the number of resistant strains to non-beta-lactam antibiotics (erythromycin, clindamycin, tetracycline, co-trimoxazole) was higher in penicillin-resistant strains compared with penicillin susceptible isolates. All isolates were susceptible to vancomycin.     

Resistance to antibiotics of Shigella strains isolated in Somalia

The resistance to antibiotics of 240 Shigella strains isolated in Somalia from 1973 to 1976 was studied. Many strains, particularly those of Shigella dysenteriae type 1, were found to be resistant to more than one drug. In view of their resistance to ampicillin, chloramphenicol, streptomycin, tetracycline, and sulfonamides, it is suggested that polymyxin B or M sulfate - which have proved to be effective in vivo - should be used for the treatment of clinically typical cases of bacillary dysentery.     

Effects of the major Pasteurella multocida porin on bovine neutrophils

OBJECTIVE: To evaluate in vitro effect of the major fraction of outer membrane proteins of Pasteurella multocida with porin-like activities on some biological functions of bovine neutrophils. ANIMALS: Neutrophils from 5 adult cattle. PROCEDURE: Variations in such biological processes as actin polymerization and chemotaxis and evaluation of hydrogen peroxide attributable to variable concentrations of P multocida were recorded and compared. Data were obtained, using the porin and lipopolysaccharide (LPS) isolated from a strain of P multocida cultivated in brain-heart infusion (BHI) broth. Various concentrations of porin and LPS were analyzed to evaluate changes in functional activation and microbicidal activity of bovine neutrophils. RESULTS: The 37.5-kd major polypeptide of the outer membrane of P multocida was isolated. Presence of this porin was significantly correlated with variations of some biological functions of bovine neutrophils. These immunocompetent cells had a concentration-dependent increase in actin polymerization and chemotactic activity. A concentration-dependent variation in the oxidative burst also was observed. CONCLUSIONS: The porins of gram-negative bacteria affect several biological functions of cells involved in the immune response as well as in inflammation. Significant correlation of results of in vitro experiments also was identified between porin and LPS effect. Pretreatment of bovine neutrophils with various concentrations of porin always caused a concentration-dependent increase in examined biological activities.     

Virulence of capsulated and noncapsulated isolates of Pasteurella multocida and their adherence to porcine respiratory tract cells and mucus

The virulence and the adherence to porcine respiratory tract cells and mucus of three toxigenic, capsular type D Pasteurella multocida isolates and their noncapsulated variants were evaluated in the present study. Loss of capsule by P. multocida, verified by transmission electron microscopy after polycationic ferritin labeling, was associated with a massive reduction in virulence of the organisms in mice. Specific-pathogen-free piglets inoculated intranasally with one of the capsulated isolates or its noncapsulated variant developed turbinate lesions characterized by bone resorption and by an inflammation of the mucosa associated with hyperplasia and squamous metaplasia of the epithelium. Infection with the capsulated isolate led to more severe lesions and atrophy of turbinates. The interactions of these P. multocida isolates with porcine respiratory tract cells and mucus were studied in vitro. The presence of capsule resulted in a decrease in binding of respiratory tract mucus were studied in vitro. The presence of capsule resulted in a decrease in binding of respiratory tract mucus to P. multocida isolates as determined by a dot blot assay. The presence of capsule also resulted in a significant decrease in adherence to porcine tracheal rings maintained in culture. The capsule seemed to mask outer membrane components which are involved in adherence. One of these components might be lipopolysaccharide since purified lipopolysaccharide bound respiratory tract mucus and blocked adherence of this microorganism to porcine tracheal rings. Our data indicate that capsular material does not seem to be involved in adherence of P. multocida to respiratory tract cells and mucus, but capsulated isolates are more virulent in mice and also in piglets.     

In vivo effect of Pasteurella haemolytica infection on bovine neutrophil morphology

OBJECTIVE: To determine whether characteristic changes in neutrophil morphology caused in vitro by Pasteurella haemolytica leukotoxin (LKT) can be observed in vivo by electron microscopic examination of infected tissue chamber fluids and pneumonic lungs. ANIMALS: 7 mixed-breed beef calves. PROCEDURE: Tissue chambers were implanted subcutaneously in 3 calves and were inoculated with P haemolytica or phosphate-buffered saline solution. Chamber fluid samples, obtained at 8 and 32 hours after inoculation, were examined, using electron microscopy. Experimental pneumonia was induced in an additional 4 calves by transthoracic inoculation with P haemolytica. These calves were euthanatized at 6, 12, 24, and 36 hours after inoculation and lung sections were examined, using transmission electron microscopy. RESULTS: On examination, using transmission electron microscopy, neutrophils in lung sections and tissue chamber fluids had cytoplasmic and nuclear changes indicative of irreversible cell injury, including cell swelling, loss of plasma membrane ruffling, mitochondrial swelling, autolytic vacuolation, disruption of plasma membrane, nuclear pyknosis, karyolysis, and karyorrhexis. On examination, using scanning electron microscopy, leukocytes obtained from tissue chambers did not have their typical convoluted surfaces, but appeared rounded and swollen or shrunken with pitted surfaces. CONCLUSIONS: Pasteurella haemolytica-induced changes in neutrophil morphology in vivo were similar to those previously induced by in vitro exposure of neutrophils to LKT. Changes were suggestive of injury initiated by damage to the plasma membrane, which is consistent with the mechanism of action of pore-forming cytolysins. CLINICAL RELEVANCE: Pasteurella haemolytica LKT appears to be an important virulence factor in vivo; a fact that should be addressed in the development of vaccines.     

Transposons coding beta-lactamase with an extended spectrum of effects and resistance to other antibiotics

The authors present contemporary knowledge concerning the transposition of resistance genes to penicillins, cephalosporins and carbapenems, and transposons and integrins coding resistance to other antibacterial substances. Transposition is, together with bacterial conjugation and transduction with bacteriophages, another mechanism of mobility and restructuring of resistance genes in bacterial strains of the same and also in other bacteria.     

Effects of Pasteurella multocida toxin on porcine bone marrow cell differentiation into osteoclasts and osteoblasts

The effect of Pasteurella multocida toxin (PMT) on porcine osteoclast and osteoblast differentiation was studied using in vitro cell culture systems. When grown in the presence of Vitamin D3, isolated porcine bone marrow cells formed multinucleated cells with features characteristic of osteoclasts. Exposure of bone marrow cells to Vitamin D3 and PMT during growth resulted in formation of increased numbers and earlier appearance of osteoclasts compared to controls. Ultrafiltered medium form PMT-treated cells likewise increased osteoclast numbers, suggesting that a soluble mediator may be involved in the action of PMT. When cell cultures were treated with fluorescein-labeled PMT, fluorescence was found within the cytoplasm of small, round cells that did not resemble either osteoclasts or osteoclastic precursor cells. Cultures of porcine bone marrow cells exposed to dexamethasone, ascorbic acid, and beta-glycerophosphate developed into osteoblastic cells that formed multilayered, mineralized nodules. Exposure of osteoblastic cultures to low concentration of PMT resulted in retarded cell growth, formation of decreased numbers of nodules and minimal to no mineralization in the nodules; higher concentration of PMT resulted in increased cellular debris and poor growth of cells, with no nodule formation. These findings suggest that PMT may induce turbinate atrophy in pigs by increasing osteoclast numbers and inhibiting osteoblastic bone formation. The effect of PMT on osteoclastic differentiation and growth may not be due to a direct effect on preosteoclastic cells, but rather due to alterations in the soluble mediator secretion by marrow stromal cells.     

Characterization of monoclonal antibodies generated against bovine and porcine prostacyclin synthase and quantitation of bovine prostacyclin synthase

Monoclonal antibodies were raised against prostacyclin synthases purified from bovine and porcine aortae, respectively. Two monoclonal antibodies, RS1 and RS2, were purified and characterized. As shown by enzyme activity precipitation and Western blot analysis, in solubilized bovine and porcine aortae microsomes the monoclonal antibodies reacted only with prostacyclin synthase. The monoclonal antibody RS1 cross-reacts with partially purified prostacyclin synthase from human umbilical veins in an ELISA-based assay. None of the antibodies inhibited the enzyme activity. By combination of the monoclonal antibody RS2 with a polyclonal antibody we established an enzyme-linked immunosorbent assay (ELISA) for quantitation of bovine prostacyclin synthase. ELISA data were confirmed by Western blot analysis. Among different bovine tissues, aortae with 1665 +/- 200 ng/mg microsomal protein showed the highest content of PGIS. Significant lower concentrations were observed in tongue, lung, kidney and thymus ranging from 49 +/- 13.4 to 2.7 +/- 0.9 ng/mg protein. The monoclonal antibody RS1 binds to endothelial cells and vascular smooth muscle cells in human liver tissue.     

Catecholaminergic region A15 in the bovine and porcine hypothalamus

Magnocellular perikarya within the retrochiasmatic division of the supraoptic nucleus of bovine and porcine hypothalami were immunoreactive (ir) with antiserum against tyrosine hydroxylase (TH), but not dopamine-beta-hydroxylase (DBH). Few cells in this region were also immunoreactive for vasopressin (VP) or oxytocin (OT). In contrast, the main division of the supraoptic nucleus contained numerous perikarya immunoreactive for VP and OT, but not TH nor DBH. Both the retrochiasmatic and principal divisions of the supraoptic nuclei contained TH- and DBH-ir fibers and varicosities. This region in bovine and porcine hypothalami corresponds to the ventral A15 catecholaminergic (dopamine-producing) cell group.     

UDP-glucose dehydrogenase from bovine liver: primary structure and relationship to other dehydrogenases

The primary structure of bovine liver UDP-glucose dehydrogenase (UDPGDH), a hexameric, NAD(+)-linked enzyme, has been determined at the protein level. The 52-kDa subunits are composed of 468 amino acid residues, with a free N-terminus and a Ser/Asn microhetergeneity at one position. The sequence shares 29.6% positional identity with GDP-mannose dehydrogenase from Pseudomonas, confirming a similarity earlier noted between active site peptides. This degree of similarity is comparable to the 31.1% identity vs. the UDPGDH from type A Streptococcus. Database searching also revealed similarities to a hypothetical sequence from Salmonella typhimurium and to "UDP-N-acetyl-mannosaminuronic acid dehydrogenase" from Escherichia coli. Pairwise identities between bovine UDPGDH and each of these sequences were all in the range of approximately 26-34%. Multiple alignment of all 5 sequences indicates common ancestry for these 4-electron-transferring enzymes. There are 27 strictly conserved residues, including a cysteine residue at position 275, earlier identified by chemical modification as the expected catalytic residue of the second half-reaction (conversion of UDP-aldehydoglucose to UDP-glucuronic acid), and 2 lysine residues, at positions 219 and 338, one of which may be the expected catalytic residue for the first half-reaction (conversion of UDP-glucose to UDP-aldehydoglucose). A GXGXXG pattern characteristic of the coenzyme-binding fold is found at positions 11-16, close to the N-terminus as with "short-chain" alcohol dehydrogenases.(ABSTRACT TRUNCATED AT 250 WORDS)     

Modification of vancomycin nephrotoxicity by other antibiotics in rats

Vancomycin (VCM) was intravenously administered to rats for 14 days at doses of 150 mg/kg/day and 250 mg/kg/day alone or in combination with 1,000 mg/kg/day of latamoxef (LMOX), flomoxef (FMOX) or cefpirome (CPR) or 250 mg/kg/day of fosfomycin (FOM), and the influences of combined antibiotics on the VCM-induced renal damage were studied. The renal impairment caused by VCM alone was, morphologically, demonstrated mainly as regeneration of tubular epithelium: slight regeneration was observed in a half of rats administered 150 mg/kg/day and slight to extensive regeneration in all the rats administered 250 mg/kg/day. Clinical examinations found apparent increases in urinary LDH and MDH activities in rats administered 250 mg/kg/day, thus showing a good correlation with renal pathological changes. In addition, increase in kidney weight and increase in urinary NAG activity were noted, while changes in plasma urea-N and creatinine were mild, and gamma-GTP activity and protein in urine could not be used as a parameter of the renal impairment. The slight renal impairment as noted in rats administered VCM 150 mg/kg/day alone was not observed at all when LMOX or FMOX was administered concomitantly, and less pronounced even when FOM was administered concomitantly. When CPR was administered concomitantly, the changes were the same as those observed with VCM alone. The renal impairment in rats administered VCM 250 mg/kg/day was apparently less severe when combined with LMOX, FMOX and FOM than that in rats administered VCM alone, and this was supported by apparent reduction of clinical examination values as the parameter of VCM-induced nephrotoxicity.(ABSTRACT TRUNCATED AT 250 WORDS)     

Determination of tilmicosin in bovine and porcine sera by liquid chromatography

A liquid chromatographic (LC) assay is described for determining tilmicosin in bovine and porcine blood sera. Tilmicosin is isolated from the serum matrix and purified by solid-phase extraction with C18 sorbent. Sample is analyzed by LC using a gradient system with a phenyl reversed-phase column that separates tilmicosin from the matrix in 30 min. Tilmicosin is measured by UV absorbance at 280 nm. Validation of assay included evaluation of accuracy, precision, linearity, specificity, sensitivity, range, and sample stability. The method has a limit of quantitation of 0.1 ppm and a validated range of 0.1 to 10.0 ppm. Recoveries were 91-95% for bovine serum and 85-93% porcine serum. The limit of detection was 0.05 microgram/mL. Limits of detection and quantitation were based on 3 and 6 times the baseline noise of control serum samples, respectively. Relative standard deviations of precision samples (n = 6) were 2% or less for both sera. The method has better specificity and analysis time than previous microbiological methods for tilmicosin in sera.     

Thermodynamic stability of bovine alpha-crystallin in its interactions with other bovine crystallins

Light scattering measurements were performed on dilute solutions of alpha-crystallin mixed with different combinations of beta H, beta L and gamma-fractions of bovine lens crystallins. Light scattering intensities were obtained as a function of scattering angle, concentration and temperature. The temperature dependence of the second virial coefficients was used to obtain partial molar enthalpy and end entropy of solutions. The difference between the thermodynamic parameters of the crystallin mixtures and those of the weighted averages of the individual components yielded the excess enthalpy and entropy functions of the solutions. Both the excess enthalpy and entropy functions indicated that thermodynamic stability of alpha-crystallin is progressively enhanced by its interactions with gamma [symbol: see text] (beta H + gamma) [symbol: see text] (beta H + beta L + gamma) crystallins. The last two combinations showed negative values both for excess enthalpy as well for excess entropy of solutions. Other combinations demonstrated increasing positive values. This implies that the combination of all four crystallins in the vertebrate lens enables the best solvation property as well as the best packing as opposed to any other single or combinatorial arrangements of crystallins. Similar conclusions have been obtained in the past from water and other vapor sorption studies.     

Identification of alpha-carboxamidated and carboxy-terminal glycine forms of peptides in bovine hypothalamus, bovine pituitary and porcine heart extracts

Two chemical assays have been developed for identifying and quantifying peptides which either could be biologically active by virtue of their alpha-carboxamidation or could be substrates for peptidylglycine alpha-amidating mono-oxygenase. The first assay is specific for the alpha-carboxamide of peptides. Using bis[trifluoroacetoxy]iodobenzene, the alpha-carboxamide was converted via a Hoffman reaction into a primary amine, which was then quantified by ninhydrin. The second assay is specific for glycine at the carboxy-terminus of a peptide. Glycine at the carboxy-terminus was derivatized to form 2-thiohydantoin, which was then separated and quantified by reverse phase HPLC. These assays were used to detect peptides in HPLC-separated extracts of bovine hypothalamus, bovine anterior lobe pituitary and porcine heart which may be of biological interest.     

Resistance to macrolide antibiotics found in methicillin-resistant Japanese clinical isolates of Staphylococcus aureus in 1996

OBJECTIVE: To study the effect of sufficient energy intake, by means of the protocolized administration of naso-gastric tube feeding, on the nutritional status of a child with cancer. DESIGN: A comparative experimental study. SETTING: Tertiary care at the Centre for Pediatric Oncology, South East Netherlands, University Hospital, Nijmegen. SUBJECTS: Seven children, newly diagnosed with cancer, were included in the experimental study and all completed the trial period. Fourteen patients were included in the retrospective study. They were randomly chosen from a group of patients previously treated for a malignancy at our department and who had received naso-gastric tube feeding for at least 16 weeks. INTERVENTION: Protocolized (experimental group) vs non-protocolized (retrospective group) administration of naso-gastric tube feeding over a period of 16 weeks. The main difference was the amount of tube feeding administered. In addition to energy from other foods, children in the experimental group received 106+/-13% of their total daily energy requirements (TDER) by means of tube feeding, whereas children in the retrospective group had received 75+/-24%. MAIN OUTCOME MEASURES: Weight as a percentage of weight for height according to the 50th percentile of a healthy reference population=ideal weight. RESULTS: Weight, expressed as a percentage of the ideal weight, increased significantly in the experimental group (18.2 8.4; P=0.01) and the retrospective study group (5.2 7.3; P=0.001). However, the increase was statistically significant in favour of the experimental group (P=0.003), in which all the children reached their ideal weight, compared to 21% in the retrospective group. CONCLUSION: Aggressive protocolized nutritional intervention during the intensive phase of anti-cancer treatment, in the form of naso-gastric tube feeding that provides the child's total daily energy requirements, results in considerable improvement in the nutritional status.