The diagnosis of small bowel bacterial overgrowth. Reliability of jejunal culture and inadequacy of breath hydrogen testing.

The reliability of a single jejunal culture in the diagnosis of small bowel bacterial overgrowth has recently been questioned. Seventy-seven patients thought to have bacterial overgrowth, defined as a jejunal culture yielding at least 10(6) organisms per milliliter of aspirate, took part in the study. Bacterial overgrowth was found in 74% of the patients with predisposing conditions and in 32% of those with no clear causes of bacterial colonization. The intestinal juice of some patients was taken at two different levels of the proximal jejunum, using both the closed- and open-tube systems. Highly significant correlations (rs = 0.90, p less than 0.001) were found between the numbers of bacteria per milliliter at the 2 jejunal levels and between the numbers of bacteria per milliliter of jejunal aspirate obtained from the closed and open tubes (rs = 0.84, p less than 0.001). Compared with the jejunal culture, the gas chromatography of volatile fatty acids in jejunal aspirate and the glucose- and lactulose-hydrogen breath tests showed sensitivities of 56%, 62%, and 68% and specificities of 100%, 83%, and 44%, respectively. This work demonstrates the reliability of jejunal cultures and the inadequacy of breath hydrogen testing in the prediction of positive jejunal cultures. When results of testing for volatile fatty acids in jejunal aspirates are positive, this always indicates the presence of bacterial overgrowth; thus, this procedure would avoid the more complicated, time-consuming, and costly bacteriological analysis of jejunal samples.

A new technique for segmental jejunal perfusion in man.

A multichannel tube with two occluding balloons was developed for measurements of the secretion of different endogenous substances into a defined segment of the small intestine. The tube was inserted under fluoroscopic guidance with the aid of a guide wire and was in position in the proximal jejunum after 45-90 min. The overall success rate of the complete procedure was 90%. The system prevented contamination of the perfusate from gastric and pancreatic secretion. Major dilution of the intestinal secretion was avoided due to high recovery of the perfusion fluid. The jejunal secretion rates of high and low molecular weight substances, i.e., albumin, beta 2-microglobulin, hyaluronan, and histamine, were tested. The recovery of the effluent fluid and the volume marker 14C-PEG 4000 exceeded 95% and was also stable over the perfusion period. The mean secretion rates in 23 healthy controls of albumin (635 micrograms/cm/h), beta 2-microglobulin (0.87 microgram/cm/h), hyaluronan (1061 ng/cm/h), and histamine (70 ng/cm/h) were also stable during the perfusion period. Ten healthy controls were studied under steady state conditions for 3 h. The concentrations in jejunal perfusion fluid relative to serum levels were for albumin 0.06%, indicating the degree of passive leakage from the plasma compartment to the intestinal lumen. The appearance of the low-molecular weight protein beta 2-microglobulin in perfusion fluid was on average 3% of its circulating levels, suggesting that the concentrative transport from plasma is dependent on the molecular size. The jejunal fluid concentrations of histamine and hyaluronan exceeded their respective concentrations in plasma, indicating that local intestinal secretion/synthesis is mainly responsible for their appearance in jejunal fluid. In summary, the technique is simple, rapid, atraumatic, safe, and reproducible and, potentially, can accurately reflect biochemical processes in the small intestine.

Effectiveness of pH measurements in predicting feeding tube placement.

The extent to which pH values of aspirates from feeding tubes could be used to differentiate between (a) gastric and intestinal placement, and (b) gastric and respiratory placement were determined in a clinical study. The sample consisted of 181 adult subjects, 94 with small-bore nasogastric tubes and 87 with nasointestinal tubes. Data were collected at the time of initial tube placement and again, when possible, after one or two days of tube feedings. Using color coded pH-paper, a total of 247 readings were made concurrently with x-rays to determine feeding tube position. Findings indicated that pH readings were often effective in differentiating between gastric and intestinal placement (p less than .0001). For example, approximately 81% of the aspirates from nasogastric tubes had pH values ranging from 1 through 4, while almost 88% of the aspirates from nasointestinal tubes had pH values of 6 or greater. Only one aspirate from a tube inadvertently placed in the lung was tested; as expected it had an alkaline pH.

Comparison of the reactivities and immunogenicities of procholeragenoid and the B subunit of cholera toxin in Thai volunteers.

Procholeragenoid, a stable high-molecular-weight aggregate of cholera toxin derived by heat treatment, was evaluated for reactivity and immunogenicity in adult Thai volunteers. Since procholeragenoid is known to retain some residual activity of cholera toxin, increasing amounts were ingested until diarrhea occurred; 250 micrograms induced diarrhea, but 100 micrograms did not. Procholeragenoid and cholera toxin B subunit, both in 100-micrograms amounts, were then compared for systemic and intestinal antitoxin responses. When three peroral doses were given, these immunogens gave comparable responses. The secretory immunoglobulin A antitoxin responses to three doses of 100 micrograms of B subunit did not differ significantly from responses found in previous studies of Thai adults given 1 or 5 mg of B subunit, but serum antitoxin responses were less after 1 or 2 doses of 100 micrograms than after doses of 1 or 5 mg. Serum antitoxin levels were similar after 3 doses of B subunit. Procholeragenoid in the maximum safe dose of 100 micrograms does not offer any immunologic advantage over B subunit, although it may be less expensive and easier to produce. However, these studies suggest that higher amounts of B subunit are more immunogenic and may be preferable, if found to be sufficiently cost effective, when added to oral killed whole Vibrio cholerae vaccines.

Transport of serum IgA into murine respiratory secretions and its implications for immunization strategies.

Both nonlabeled and radiolabeled IgA mAb with specificity toward Sendai virus, a respiratory pathogen, were used to investigate the transport of serum polymeric and monomeric IgA into murine upper and lower respiratory secretions as well as into the gut. After purification by affinity chromatography, IgA mAb were fractionated into monomers and polymers by gel filtration and radiolabeled with 125I. Mice were injected i.v. with either 125I-monomer and 131I-albumin or 125I-polymer and 161I-albumin. At various times after injection, serum and gut, nasal and bronchoalveolar lavage samples were collected. The TCA precipitable radioactivities were determined and the selective transport indices calculated. The results indicated selective transport of polymeric IgA but not monomeric IgA from serum into upper respiratory and intestinal secretions. The degree of TCA precipitability in nasal lavage and to a lesser extent gut secretions suggested significant degradation of the antibody during or after transport. To investigate further the integrity of the IgA in mucosal secretions, ELISA viral binding activity of nonradiolabeled IgA was determined for both IgA incubated with nasal secretions in vitro and polymeric IgA recovered by nasal lavage 4 h after i.v. injection. Although reconstitution experiments indicated no significant loss of antibody binding activity after incubation of antibody with lavage fluid in vitro, only negligible ELISA binding activity was detected in nasal washes after i.v. injection of antibody. The data overall suggest that although there is a quantitatively small, but selective transport of polymeric IgA into the upper respiratory tract, this transport results in minimal functional antibody activity. Implications of these and other findings for strategies of oral immunization in prophylaxis against respiratory infections are discussed.

Hepatobiliary scintigraphy and the string test in the evaluation of neonatal cholestasis.

We evaluated [99mTc]diisopropylphenyl-carbamoylmethylimidodiacetic acid ([99mTc]DISIDA) cholescintigraphy with measurement of duodenal fluid radioactivity collected by the string test in patients with neonatal cholestasis. Twenty-six infants with prolonged jaundice and acholic stools were studied prospectively. Twelve patients had neonatal hepatitis, 12 biliary atresia, and one each Alagille syndrome and alpha 1-antitrypsin deficiency liver disease. All infants except the biliary atresia patients and four of the neonatal hepatitis patients revealed bowel activity on scan 6 h after tracer administration. At 24 h, three of these latter patients with neonatal hepatitis and two of the patients with biliary atresia revealed bowel activity. String radioactive counts for neonatal hepatitis ranged from 99,574 to 967,205 cpm (374,504 +/- 232,210 cpm; mean +/- SD) and for biliary atresia from 8,342 to 370,346 cpm (117,149 +/- 98,698 cpm; mean +/- SD). While neither test alone was capable of correctly differentiating among all patients, those patients with biliary atresia had either a negative hepatobiliary scan at 24 h or string radioactive count below 197,007 cpm. Disparity between the hepatobiliary scan and the string radioactive counts mandates further diagnostic investigation. These data suggest that simultaneous administration of the string test with hepatobiliary scintigraphy is advantageous in the evaluation of infants with cholestatic jaundice.

Single step affinity chromatographic purification of human alpha-amylase from aspirated duodenal juice and its application in the measurement of pancreatic alpha-amylase synthesis rates in man.

Human alpha-amylase was purified from aspirated duodenal juice to electrophoretic homogeneity in a single step by affinity chromatography with the competitive inhibitor acarbose (IC50 = 1.22 mumol/l) as ligand. Duodenal juice was applied to an agarose resin to which acarbose had been coupled covalently via a 1.9 nm spacer group. Pure alpha-amylase, eluted with free acarbose, had a molecular mass of 55,000, and isoelectrofocusing revealed the presence of six isozymes with pI values of 7.3, 6.8, 6.7, 6.5, 6.4 and 6.3, all of which possessed amylase activity based on positive starch/iodine staining. The potential usefulness of this one-step purification procedure in the measurement of pancreatic alpha-amylase synthesis rates was evaluated in two control patients with non-pancreatic disease. Aspirated duodenal juice was obtained during a pulse/continuous intravenous 4 h infusion of [14C]leucine together with secretin and pancreozymin, and alpha-amylase purified using our protocol. Pancreatic alpha-amylase synthesis rates were determined from the rate of incorporation of [14C]leucine into alpha-amylase; values of 4.4 and 5.1 h were obtained for the two control patients.

[Determination of lactoferrin in the diagnosis of diseases of the pancreas]. / Stanovení laktoferinu v diagnostice onemocnení slinivky brisní.

In 29 patients with chronic pancreatitis, 15 patients with malignant tumours of the pancreas and in 30 controls lactoferrin in fluid aspirated from the duodenum was assessed during the cholecystokinin-secretin (CCK-S) test. As compared with the control group, its concentration is significantly higher in patients with chronic pancreatitis, but not in patients with malignant tumours of the pancreas. It is probable that estimation of lactoferrin in the CCK-S test may prove helpful in the differential diagnosis of pancreatic disease.

[Ph measurement in the stomach and duodenum through endoscopy]. / Endoscopon keresztül végzett pH mérés a gyomorban és a duodenumban.

The pH values were measured by means of a flexible glass-electrode through endoscope in the stomach and duodenum. The measuring instrument and technique are described. The luminal and juxtamucosal pH values were determined. The latter indicates the pH value prevailing on the epithelial surface of the viscid mucous layer. The gradient between the two values expresses the hydrogen-ion neutralizing capacity of the viscid mucus on the surface of the mucosa. Quantitative data on the first protective line of the gastric and duodenal mucous membrane may be obtained by measuring it. The pH values at different points of the stomach and duodenum were determined through endoscope in 24 patients with duodenal ulcer and in 15 controls. No significant difference was found between the two groups concerning the gastric luminal pH values, however the luminal pH of the duodenal bulb was significantly lower in patients with duodenal ulcer than that of the controls. In patients with duodenal ulcer the juxtamucosal pH value was significantly lower at several points in the stomach and in the duodenum too as compared to the controls. This refers to the insufficient neutralizing capacity of the viscid mucous layer. The pH measurement performed through endoscope furnishes in a simple way valuable clinical data, it is furthermore the first quantitative method which can be used routine-like for the examination of the barrier function of the viscid mucous surface in the stomach and duodenum of man.

Human immune response to Vibrio cholerae O1 whole cells and isolated outer membrane antigens.

The serum immunoglobulin G (IgG) and mucosal secretory IgA (SIgA) response of human volunteers challenged with Vibrio cholerae O1 was analyzed for reactivity to V. cholerae O1 antigens by the immunoblot technique. Components of both in vitro- and in vivo (rabbit ligated ileal loop)-grown V. cholerae O1 were separated by sodium dodecyl sulfate-urea-polyacrylamide gel electrophoresis. Postchallenge serum IgG reacted uniquely with 15 antigens and with greater intensity than did prechallenge serum with at least 16 antigens. Serum IgG and SIgA reacted with antigens present in preparations from the homologous challenge strain of V. cholerae as well as antigens from strains of heterologous biotype or serotype. These heterologous antigens may represent antigens responsible for protection to rechallenge with a heterologous strain of V. cholerae. All the antigens detected by postchallenge jejunal fluid SIgA had an apparent molecular size of less than 25 kilodaltons. Serum IgG and jejunal fluid SIgA also reacted with antigens unique to in vivo-grown cells and several antigens in outer membrane preparations, suggesting that studies of protective immunity and V. cholerae O1 pathogenesis should include examination of both in vitro- and in vivo-grown V. cholerae O1 cellular antigens.

Relationship of jejunostomy and urine polyamine content to refeeding and intestinal structure and function.

The polyamine content of the intestinal mucosa is increased with mucosal proliferation. Our purpose was to determine if urine and jejunostomy polyamine content increases after refeeding and correlates with intestinal length and carbohydrate absorption after intestinal resection. Five patients with intestinal remnant lengths ranging from 1.5 to 8 feet after intestinal resection and jejunostomy formation were studied after refeeding of an elemental diet. Mean putrescine and spermidine levels in the jejunostomy effluent but not in the urine increased significantly after refeeding (p less than 0.05). These changes correlated with intestinal remnant length and carbohydrate absorption. Carbohydrate absorption correlated with polyamine levels in a single patient studied longitudinally. The polyamine content of intestinal fluid may serve as a marker of the status of the intestinal mucosa and may prove useful in the dietary management of individuals with the short bowel syndrome.

Marked secretion of pyruvate in human duodenal juice stimulated with pancreozymin or secretin.

Pyruvate and lactate in duodenal aspirates were investigated to determine whether they are excreted from human pancreas as substrates for alkaline secretion as is bicarbonate. Secretion of these acids was compared with that of another organic acid, citrate, which is thought to be excreted in close relationship to digestive enzymes. All acids were assayed in the fluid obtained from 11 subjects without pancreatic diseases, before and after sequential intravenous injections of 1 unit/kg pancreozymin and 1 unit/kg secretin. Pyruvate concentrations were markedly increased by each stimulation, especially by secretin, and the cumulative excretions of pyruvate and bicarbonate after secretin stimulation were significantly correlated among the subjects. In contrast, lactate concentrations, although high just after administration of pancreozymin, declined to a considerable extent following each injection, rather similar to those of protein or citrate. These data suggest that pyruvate may be secreted from human pancreatic duct cells similar to bicarbonate secretion through mechanisms related to alkaline secretion.

[Characteristics of the venoms and digestive secretions of Aphonopelma seemanni and Sphaerobothria hoffmanni (Araneae: Theraphosidae) of Costa Rica]. / Algunas características de los venenos y secreciones digestivas de Aphonopelma seemanni y Sphaerobothria hoffmanni (Araneae: Theraphosidae) de Costa Rica.

A comparison of some components of the venoms of two Costa Rican tarantulas, Aphonopelma seemanni (Cambridge) and Sphaerobothria hoffmanni (Karsch) by polyacrylamide gel electrophoresis shows patterns similar to those of Dugesiella hentzi (Girard), a North American tarantula. The digestive secretions have proteins that do not enter the 15% gels; thus no bands are observed. The method used by the tarantulas to consume their prey involves the action of both the venom and the digestive secretions. The percent protein, pH, proteolytic activity and hemolytic activity of venom and digestive secretions of both species were determined, and a high proteolytic activity for digestive secretions was found.

Algunas características de los venenos y secreciones digestivas de Aphonopelma seemanni y Sphaerobothria hoffmanni (Araneae: Theraphosidae) de Costa Rica / Characteristics of the venoms and digestive secretions of Aphonopelma seemanni and Sphaerobothria hoffmanni (Araneae: Theraphosidae) of Costa Rica

A comparison of some components of the venoms of two Costa Rican tarantulas, Aphonopelma seeamanni (Cambridge) and Sphaerobothria hoffmanni (Karsch) by polyacrylamide gel electrophoresis shows patterns similar to those of Dugesiella hentzi (Girard), a North American tarantula. The digestive secretions have proteins that do not enter the 15% gels; thus no bands are obsorved. The method used by the tarantulas to consume their prey involves the action of both the venom and the digestive secretions. The percent protein, pH, proteolytic activity and hemolytic activity of venom and digestive secretions of both species were determined, and a high proteolytic activity for digestive secretions was found.

Rabbit intestinal and colonic mucins: isolation, partial characterization, and measurement of secretion using an enzyme-linked immunoassay.

Mucin was purified separately from the upper, mid, and distal small intestine and the proximal colon of the rabbit. The carbohydrate profiles of the three intestinal mucins were the same but differed from that of the colonic mucin, which contained less N-acetylgalactosamine but more sialic acid. All four mucins had similar polymeric structures composed of large, heterogeneous glycoprotein monomers and a smaller protein of Mr approximately 120,000, held together by disulphide bonds. The three intestinal mucins, however, were more resistant to dissociation by thiol reduction or degradation by proteolysis than colonic mucin. An antibody against a pool of the purified mucins was developed in guinea pigs and used to establish an enzyme-linked immunosorbent assay (ELISA). The antibody was highly specific for rabbit intestinal and colonic mucins, showing no cross-reactivity with nonmucin components of rabbit intestinal and colonic tissue, and very little or no reactivity with purified intestinal mucins from other species. The four purified rabbit mucins had the same affinity for the antibody and similar numbers of antigenic determinants. Antigenicity was apparently associated with the protein moeity of the mucins and was critically dependent on three-dimensional conformation, since proteolysis decreased the number of antigenic determinants while thiol reduction abolished antigen-antibody affinity. Using the ELISA, the tissue mucin content and the rate of mucin secretion were found to be significantly higher in the proximal colon that in the three regions of the intestine, which were the same.