Synergistic effect of nikkomycin Z with caspofungin and micafungin against Candida albicans and Candida parapsilosis biofilms.

Antifungal lock therapy has received significant interest in the last few years because the frequently usage of intravascular devices is associated with an increasing number of catheter-related bloodstream infections caused by Candida species. Antifungal combinations with synergistic interaction can be a good choice for antifungal lock therapy; therefore, interactions were examined between two echinocandins (caspofungin and micafungin) and the chitin synthesis inhibitor nikkomycin Z against Candida albicans and C. parapsilosis biofilms. Susceptibility was evaluated using the XTT-based checkerboard microdilution method, while the nature of interactions was assessed by calculating fractional inhibitory concentration indices and using the Bliss independence model. Mathematic-based evaluations were supplemented with fluorescent LIVE/DEAD viability assay. The results obtained by statistical interaction analyses correlated well with the viability assay. The tested echinocandins with nikkomycin Z caused an extended cell death and the structure of the biofilm was sparse compared to the control, especially for C. albicans. The findings support the simultaneous usage of nikkomycin Z and caspofungin or micafungin in alternative therapies such as the antifungal lock therapy. SIGNIFICANCE AND IMPACT OF THE STUDY: Antifungal lock therapy can be a potential therapeutic approach to eradicate the intraluminal Candida biofilms; however, there is no approved lock strategy against fungal species so far. The results of this study provide valuable evidence that nikkomycin Z acts synergistically in combination with caspofungin or micafungin against biofilms. In addition, this synergy was more pronounced for micafungin combined with nikkomycin Z. Therefore, nikkomycin Z can be considered as a potential agent in antifungal lock therapy especially with micafungin against C. albicans or C. parapsilosis biofilms.

Predictors of relapse in patients with Crohn's disease in remission after 1 year of biological therapy.

BACKGROUND: Some of the most important questions relating to the use of biological therapy in inflammatory bowel diseases concern the duration of maintenance therapy. AIM: To assess the disease course and frequency of relapse of Crohn's disease (CD) following discontinuation of biological therapy, and to determine predictive factors for relapse. METHODS: One hundred twenty-one CD patients who had achieved clinical remission following 1 year of biological therapy and for whom biological therapy was then discontinued participated in this prospective observational study. Eighty-seven CD patients had received infliximab and 34 adalimumab. The definition of relapse was an increase of >100 points in CDAI to at least a CDAI of 150 points. RESULTS: Biological therapy was restarted within 1 year of treatment cessation in 45% of patients. Logistic regression analysis revealed that previous biological therapy (P = 0.011) and dose intensification during the 1-year course of biological therapy (P = 0.024) were associated with the need for and the time to the restarting of biological therapy. Smoking was observed to have an effect that was not statistically significant (P = 0.053). CONCLUSIONS: Biological therapy was restarted a median of 6 months after discontinuation in almost half of Crohn's disease patients in who had been in clinical remission following 1 year of biological therapy. These results suggest that, in the event of the presence of certain predictive factors, biological therapy should probably be continued for more than 1 year by most patients.

Water enema CT examination of rectum cancer by reduced amount of water.

OBJECTIVE: To define whether volume of water, administered during water enema CT (WE-CT) for local staging of rectal cancer, may be reduced without compromising the diagnostic value of the examination. MATERIALS AND METHODS: 29 patients with rectum cancer underwent preoperative WE-CT. Contrast-enhanced CT (equilibrium phase) measurements were performed after i.v. injection of smooth muscle relaxant and rectal administration of 400-500 ml lukewarm tap water. Quality of the obtained scans was evaluated and the images were analyzed for depth of tumor invasion. Results of the CT examinations were compared to findings at surgery. RESULTS: Despite reduced dose of water enema, 19/29 examinations were of excellent quality, 6/29 good, and 4/29 poor, but still diagnostic. We achieved sensitivity (90.1), specificity (70.1) and accuracy (86.2) in differentiating tumors confined to the bowel wall from those extending beyond it. CONCLUSION: Large volume of water enema administered during CT examination of the rectum may cause complaints and increases the risk of complications. Our results prove that using lower amount of water does not impair the quality of examination and accuracy of local staging of rectum carcinomas.

The ursodeoxycholic acid-p-aminobenzoic acid deconjugation test, a new tool for the diagnosis of bacterial overgrowth syndrome.

OBJECTIVE: To determine the possible complementary role of the ursodeoxycholic acid-p-aminobenzoic acid (UDCA-PABA) loading test in the diagnosis of intestinal bacterial overgrowth. DESIGN: A prospective clinical study. PATIENTS AND METHODS: The hydrogen breath and UDCA-PABA tests were performed simultaneously in 68 patients with suspected contaminated small bowel syndrome (CSBS), and in 10 healthy control subjects. The hydrogen breath test was performed by oral loading of 25 g of lactose and/or 10 g of lactulose. The UDCA-PABA test was carried out by oral loading of 250 mg of UDCA-PABA conjugate, followed by measurement of the amount of PABA excreted in the urine. The diagnosis of bacterial overgrowth was considered to be established when either the hydrogen breath test or the UDCA-PABA test produced abnormal results. RESULTS: Thirty-five of the 68 patients proved to have CSBS. In 13 of these 35 patients, only the enhanced urinary PABA excretion (11.7 +/- 1.42 mg vs. 3.6 +/- 0.68 mg) indicated bacterial overgrowth, 15 of the 35 patients gave only a positive hydrogen breath test, and in the remaining seven cases the results of both tests were abnormal. In eight CSBS patients, the urinary excretion of PABA was decreased significantly following 10-day tinidazole treatment (5.5 +/- 1.29 mg vs. 13.1 +/- 2.07 mg). CONCLUSION: The UDCA-PABA test is a valuable clinical method for the detection of bacterial overgrowth, especially in cases where hydrogen production alone fails to reveal CSBS. It is also a useful procedure for evaluating the efficacy of antibacterial treatment.

[The ursodeoxycholic acid-p-aminobenzoic acid test in the diagnosis of small bowel bacterial overgrowth syndrome]. / Az ursodezoxikólsav-p-aminobenzoesav teszt a kontaminált vékonybélszindróma diagnosztikájában.

UNLABELLED: Contaminated small bowel syndrome is frequently associated with meteorism due to excessive gas formation, and diarrhoea as a result of bacterial fermentative processes, including splitting of carbohydrates or deconjugating and dehydroxylating bile salts. In addition to gas production, bacteria capable of metabolizing bile salts have been shown to release p-aminobenzoic acid (PABA) from and Ursodeoxycholic-acid-PABA substrate. Our aim was to determine the possible complementary role of the UDCA-PABA test in the diagnosis of bacterial overgrowth. PATIENTS AND METHODS: The H2 breath and UDCA-PABA tests were performed simultaneously on 46 patients with suspected contaminated small bowel syndrome, and on 7 healthy subjects. The H2 breath test was performed by oral loading of 25 g lactose and/or 10 g lactulose. The UDCA-PABA test was carried out by determining urinary excretion of PABA after oral loading with 250 mg UDCA-PABA conjugate. The diagnosis of bacterial overgrowth was established, when either H2 breath, or UDCA-PABA test proved to be pathological. RESULTS: Based upon the pathologic values of either the H2 breath test, or the UDCA-PABA test, 25 out of 46 patients proved to have contaminated small bowel syndrome. In 10 out of 25 patients only pathologic urinary PABA excretion (12.772 +/- 1.707 vs 4.1 +/- 0.58), indicated bacterial overgrowth, and in 9 out of the same group only positive H2 breath test (early rise of > 20 ppm of H2) indicated the same, while in 6 cases both tests proved to be pathological. In 7 CSBS patients the urinary excretion of PABA significantly decreased following a 10 day Tinidazole treatment (5.48 +/- 1.286 vs 13.068 +/- 2.068). CONCLUSION: The UDCA-PABA test proved to be a valuable complementary method to detect bacterial overgrowth, when H2 production failed to reveal bacterial overgrowth.

A rhythmic modulatory gating system in the stomatogastric nervous system of Homarus gammarus. II. Modulatory control of the pyloric CPG.

1. In the European rock lobster, Homarus gammarus, two bilaterally symmetrical pairs of commissural neurons, P and commissural pyloric (CP), evoke excitatory postsynaptic potentials in the neurons of the pyloric motor network. The present paper shows that the two commissural neurons also exert a modulatory control over the pyloric network. 2. The P and CP neurons were active during ongoing pyloric rhythms. Ongoing pyloric activity was terminated when the neurons were hyperpolarized to inhibit their firing. 3. When the pyloric network was quiescent, depolarizing either the P or CP neuron induced a robust pyloric rhythm. 4. We studied the actions of the P and CP neurons on individual pyloric neurons isolated in situ from network interactions by a photoinactivation techniques. The P neuron induced oscillatory properties in the pacemaker pyloric dilator (PD) neurons and the motor neuron, ventricular dilator (VD), whereas the CP neuron induced rhythmogenic properties in all the network neurons but VD. Together, the P-CP neurons modulated the entire pyloric network. The modulatory effects of the P-CP neurons did not outlast the duration of their discharge. 5. The P and CP neurons also controlled the firing frequency of all the pyloric neurons. They may, in addition, control phasing of the constrictor neurons discharges, but this effect was state-dependent and occurred only when the pyloric central pattern generator was functioning weakly. Their role in providing flexibility to the network operation appeared relatively limited. 6. We conclude that the P and CP neurons are good candidates for insuring long-term maintenance of pyloric network activity patterns.

A rhythmic modulatory gating system in the stomatogastric nervous system of Homarus gammarus. I. Pyloric-related neurons in the commissural ganglia.

1. Operation of the pyloric neural network in the crustacean stomatogastric ganglion (STG) depends on constant firing of modulatory inputs from anterior ganglia. We have identified two bilaterally symmetrical pairs of these inputs in the commissural ganglia (COGs) of the European rock lobster Homarus gammarus. During operation of the pyloric CPG, they fired in pyloric time, out of phase with the pyloric pacemakers. 2. One of the pair was the commissural pyloric (CP) neuron and the other was homologous to the P neuron described in the spiny lobster Panulirus interruptus. We describe their morphology and location in the COG. The CP neuron projected to the STG via the superior esophageal nerve (son) and the stomatogastric nerve (stn), whereas the P neuron projected via the inferior esophageal nerve (ion) and stn. 3. To determine the total number of commissural neurons projecting to the STG, we used cobalt and Lucifer yellow backfilling from their cut axons in the stn. With the ion cut, there were between 8 to 12 labeled somata in each COG including CP cell body, whereas only 2 somata (including P) were labeled with the son cut. Among these neurons, CP and P appeared to be the only commissural neurons that fired in pyloric time and projected in the STG on the pyloric network. 4. The CP neuron produced monosynaptic excitatory postsynaptic potentials (EPSPs) on the pyloric dilator (PD), lateral pyloric (LP), and inferior cardiac (IC) neurons, whereas the P neuron produced monosynaptic EPSPs on all pyloric motoneurons but IC. The P neuron was gamma-aminobutyric acid immunoreactive, and the P-derived EPSPs in pyloric neurons were reversibly blocked by bicuculline, picrotoxin, and D-tubocurarine. 5. The CP and P neurons were electrically coupled, and modification of membrane potential in either one of them appreciably changed the firing frequency of the coupled neuron. 6. A negative-feedback loop from the pyloric anterior burster (AB) interneuron provoked simultaneous rhythmic inhibitions in the P and CP neurons. Together with the electrical coupling, the rhythmic inhibition contributed to synchronize firing of the two commissural neurons. 7. The following papers in the series of describe the modulatory and rhythmic control exerted by the P and CP neurons over the pyloric pattern generator.

A rhythmic modulatory gating system in the stomatogastric nervous system of Homarus gammarus. III. Rhythmic control of the pyloric CPG.

1. Two modulatory neurons, P and commissural pyloric (CP), known to be involved in the long-term maintenance of pyloric central pattern generator operation in the rock lobster Homarus gammarus, are members of the commissural pyloric oscillator (CPO), a higher-order oscillator influencing the pyloric network. 2. The CP neuron was endogenously oscillating in approximately 30% of the preparations in which its cell body was impaled. Rhythmic inhibitory feedback from the pyloric pacemaker anterior burster (AB) neuron stabilized the CP neuron's endogenous rhythm. 3. The organization of the CPO is described. Follower commissural neurons, the F cells, and the CP neuron receive a common excitatory postsynaptic potential from another commissural neuron, the large exciter (LE). When in oscillatory state, CP in turn excites the LE neuron. This positive feedback may maintain long episodes of CP oscillations. 4. The pyloric pacemaker neurons follow the CPO rhythm with variable coordination modes (i.e., 1:1, 1:2) and switch among these modes when their membrane potential is modified. The CPO inputs strongly constrain the pyloric period, which as a result may adopt only a few discrete values. This effect is based on mechanisms of entrainment between the CPO and the pyloric oscillator. 5. Pyloric constrictor neurons show differential sensitivity from the pyloric pacemaker neurons with respect to the CPO inputs. Consequently, their bursting period can be a shorter harmonic of the bursting period of the pyloric pacemakers neurons. 6. The CPO neurons seem to be the first example of modulatory gating neurons that also give timing cues to a rhythmic pattern generating network.

A receptor with GABAC-like pharmacology in invertebrate neurones in culture.

We have characterized in crustacean neurones in culture a receptor for gamma-aminobutyric acid (GABA) which conforms to the pharmacological profile of the proposed type-C GABA receptor (GABAC) found in the vertebrate retina. It is associated with a chloride-selective ion channel and is blocked by picrotoxin. It is neither inhibited by bicuculline nor activated by baclofen, while diazepam and phenobarbital are without modulatory effect. Like the GABAC-like receptor of the vertebrate retina it is activated by the folded GABA analogue cis-4-aminocrotonic acid (CACA). Desensitization is moderate allowing for a more sustained action of GABA. Single channel recordings revealed a bicuculline-resistant GABA- and CACA-activated chloride channel with a conductance about eight times higher than that described for the bicuculline-resistant GABA receptor channel from the rat retina.

Muscarinic modulation of a pattern-generating network: control of neuronal properties.

The aim of this article is to investigate the cellular mechanisms underlying cholinergic modulation of the pyloric network in the stomatogastric ganglion (STG) of the Cape lobster Jasus Ialandii. Bath application of the muscarinic agonists muscarine, oxotremorine, and pilocarpine on the STG activates a rhythmic pattern from a quiescent pyloric network. The mechanisms of this modulation were investigated on individual pyloric neurons isolated both from synaptic interactions within the network (by photoinactivation of most of the presynaptic neurons and pharmacological blockade of the remaining synapses) and from central inputs (by a sucrose block of the input nerve). All three muscarinic agonists activated bursting and plateau properties of all the neurons comprising the pyloric network. The activation was dose dependent, and was blocked by the muscarinic antagonists atropine, pirenzepine, and scopolamine. The oscillatory behavior triggered by the muscarinic stimulation was specific to each type of pyloric neuron. The isolated neuron AB had the shortest oscillation period and depolarizing phase. The constrictor neurons (LP, PY, IC) were the slowest oscillators, and only oscillated upon hyperpolarizing current injection. Under muscarinic modulation, the individual bursting activities of the isolated pyloric neurons were of the same type as their activities when isolated from the network but modulated by central inputs (Bal et al., 1988). The VD neuron is an exception since it was a rapid oscillator in the latter situation and became a slow oscillator when modulated by a single muscarinic agonist. To determine the relative importance of the muscarinic-dependent bursting properties of the individual pyloric neurons in the operation of the intact network, a progressive reconstruction of the synaptic circuitry was attempted. We found that under certain conditions of muscarinic modulation a new composite pacemaker could be created, composed of the electrically coupled VD, AB, and PD neurons. This can result in the generation of new pyloric patterns that were very sensitive to the membrane potential of individual network neurons. The data also confirmed that, in a rhythmic "pattern-generating network," the pacemaker role may not be definitely attributed to a given neuron but instead could be assigned to other neurons by modulation of their respective oscillatory capabilities.

Combination of horseradish peroxidase and lucifer yellow staining for selective labeling of neurons at the electron microscopic level.

We have developed a new double labeling method for electron microscopy to characterize selectively two physiologically identified neurons on the same preparation. The stomatogastric nervous system of crustaceans was used to test the distinguishing staining characteristics of the two labels. Neurons were labeled on one side with horseradish peroxidase (HRP) and on the other side with Lucifer yellow (LY). After blue light irradiation of the tissue in the presence of diaminobendizine, the two labeled neurons could be easily observed and discriminated on the same section by the two different reaction products. This simple technique of double labeling is useful in experimental neuroanatomy for the detailed study of synaptic relationships.

Suppressive control of the crustacean pyloric network by a pair of identified interneurons. I. Modulation of the motor pattern.

A pair of identified neuromodulatory neurons, the pyloric suppressor (PS) neurons, can individually and strongly modify the activity of the pyloric network in the stomatogastric nervous system of the lobster Homarus gammarus. The PS neurons are identified by the location of their somata in the inferior ventricular nerve, their axonal projections, and their effects on pyloric network activity in vitro. Discharge of a PS neuron evokes large EPSPs in the pyloric dilator (PD) neurons and a long-lasting cessation of rhythmic activity in the neurons that control movements of the pyloric filter: PD, lateral pyloric (LP), and pyloric (PY). This cessation of rhythmic activity can outlast by several 10s of seconds a brief discharge of PS lasting only a few seconds. The different neurons of the pyloric filter do not exhibit the same sensitivity to the suppressive effects of PS, with the LP neuron being the most sensitive. Tonic discharge in PS induces graded alterations in the pyloric pattern, depending on its firing frequency. At low (less than 5 Hz) discharge frequencies, PS provokes changes in phase relationships and duration of bursting in pyloric neurons. A slight increase in PS frequency suppresses the rhythmic activity of some pyloric neurons, resulting in a switch from a triphasic to a biphasic pattern. At higher (greater than 10 Hz) PS firing frequencies, rhythmic activity in all the pyloric neurons, including the pacemakers (PD, anterior burster), is abolished, except in cells (ventricular dilator, inferior cardiac) controlling the pyloric valve. We conclude that a central pattern generator is not only subject to activating modulatory control, but may also be the target of suppressive inputs that are themselves able to provoke functional reconfigurations of the network.

Suppressive control of the crustacean pyloric network by a pair of identified interneurons. II. Modulation of neuronal properties.

In the lobster Homarus, the 2 identified PS neurons have a strong suppressive modulatory effect on the activity of the pyloric network in the STG (Cazalets et al., 1990). In the present paper, we consider the effects of PS on individual pyloric neurons isolated from their partners in the network by cell photoinactivation and synaptic blockade. Three types of PS action are described: (1) a transient, EPSP-mediated depolarization of the PD, VD, and AB neurons; (2) a long-lasting hyperpolarization concomitant with a loss of oscillatory properties in the PD and LP neurons; (3) a long-lasting depolarization without modification of oscillatory properties in the PY and IC neurons. The various effects of PS on isolated pyloric cells were consistent with the overall effects of PS on the intact pyloric network.

Experience with topical administration of 4-aminosalicylic acid in ulcerative colitis.

4-Aminosalicylic acid was applied topically in a daily dose of 1.4 gm for two weeks in ten patients with ulcerative colitis. After favorable results, the therapeutic effects of 4-aminosalicylic acid and salazopyrin enemas were compared in a two-week cross-over open trial, in 20 patients suffering from recurrent ulcerative colitis involving the rectum and rectosigmoid. No significant difference was found in the changes of the endoscopic picture of the mucosa. The results did not show a significant difference between 4-aminosalicylic acid and salazopyrin enemas, either in the clinical activity or in the histologic picture. 4-Aminosalicylic acid seems to be a suitable drug for improving the clinical symptoms of ulcerative proctitis.

Control by an identified modulatory neuron of the sequential expression of plateau properties of, and synaptic inputs to, a neuron in a central pattern generator.

Recordings from the lateral gastric (LG) neuron, which forms part of the gastric mill central pattern generator in the red lobster, Palinurus vulgaris, indicate that regenerative membrane properties (plateau properties) and synaptic inputs interact sequentially rather than simultaneously to determine its discharge pattern. LG thus presents a composite discharge, consisting of 2 separate segments of firing and one silent period. The first firing segment depends on regenerative membrane properties; this is the endogenous component, or segment, of LG's discharge. The second firing segment is the result of extrinsic synaptic input, forming the synaptic component of LG's discharge. The relative importance of these 2 components can vary, and thus LG's discharge ranges from one in which LG fires only as a result of its endogenous component to one in which its endogenous component is entirely absent and only the synaptic component underlies action potentials. Activity in an identified modulatory neuron suppresses the endogenous segment and enhances the synaptic segment of LG's discharge. This long-lasting effect in turn changes phase relationships within the gastric mill network and provides mechanisms for producing flexibility in the gastric pattern generator and for ensuring that a specific motor output is generated by a flexible neural network.

Suppressive control of a rhythmic central pattern generator by an identified modulatory neuron in crustacea.

The activity of the 14 neuron network which organizes the pyloric motor rhythm in the stomatogastric ganglion of the lobster, Homarus gammarus, is controlled by neuromodulatory inputs which have been described as having mainly 'permissive' effects. By contrast, here we identify a neuron, the pyloric suppressor (PS) neuron which exerts a 'suppressive' effect on the pyloric activity. We show that PS neuron discharge can terminate in a long-lasting manner, spontaneous pyloric rhythmic activity. Its effect results from a direct suppression of the endogenous ability of the pyloric pacemaker neurons to produce rhythmic bursts of action potentials. Thus the output of the pyloric neuronal network appears to be finely tuned by neuromodulatory influences having opposite effects.

Control of a central pattern generator by an identified modulatory interneurone in crustacea. I. Modulation of the pyloric motor output.

In the lobsters Fasus lalandii and Palinurus vulgaris, the rhythmical activity of the pyloric pattern generator of the stomatogastric nervous system is strongly modified by the firing of a single identified interneurone, whose activity we have recorded from the cell body, in vitro. The cell body of this interneurone, the anterior pyloric modulator (APM), is located in the oesophageal ganglion and sends two axons to the stomatogastric ganglion via the inferior oesophageal nerves, the commissural ganglia, the superior oesophageal nerves and the stomatogastric nerve. Firing of neurone APM modifies the activity of all the neurones of the pyloric network, including pacemaker and follower neurones. Its effects are both quantitative (increase in the frequency of the rhythm and in the frequency of spikes within cell bursts) and qualitative (modifications in relative efficacies of the synaptic relationships within the pyloric network, which in turn lead to changes in the phase relationships between the discharges of the neurones). The effects on pyloric activity induced by firing of neurone APM are established slowly (one or two seconds) and are of long duration (ten times the duration of APM's discharge). These modifications most probably involve muscarinic cholinergic receptors. APM's influences on the activity of pyloric neurones appear to be characteristic of a neuromodulatory process and are such that they may be of behavioural significance in the intact animal.

Control of a central pattern generator by an identified modulatory interneurone in crustacea. II. Induction and modification of plateau properties in pyloric neurones.

In the isolated stomatogastric nervous system of the lobster Fasus lalandii, the strong modifications of the pyloric motor pattern induced by firing of the single anterior pyloric modulator neurone (APM) are due primarily to modulation by APM activity of the regenerative membrane properties which are responsible for the 'burstiness' of all the pyloric neurones and particularly of the non-pacemaker neurones (constrictor motoneurones). This modulation has been studied under experimental conditions where the main extrinsic influences usually received by the pyloric constrictor neurones (intra-network synaptic interactions, activity of pacemaker neurones, and phasic central inputs from two premotor centres) are minimal. Under these conditions a brief discharge of neurone APM induces long plateaus of firing in all of the pyloric neurones. The non-pacemaker neurones of the pyloric network are not simply passive follower neurones, but can produce regenerative depolarizations (plateau potentials) during which the neurones fire spikes. The ability of the pyloric constrictor neurones to produce plateau potentials (plateau properties) contributes greatly to the generation of the rhythmical pyloric motor pattern. When these neurones spontaneously express their plateau properties, firing of neurone APM amplifies these properties. When most of the central inputs usually received by the pyloric constrictor neurones are experimentally suppressed, these neurones can no longer produce plateau potentials. In such conditions, firing of the single modulatory neurone APM can reinduce plateau properties of the pyloric constrictor neurones. In addition, firing in APM neurone slows down the active repolarization phase which terminates the plateau potentials of pyloric constrictor neurones. This effect is long-lasting and voltage-dependent. Modulation by APM of the plateau properties of the pyloric neurones also changes the sensitivity of these neurones to synaptic inputs. This effect can explain the strong modifications that an APM discharge exerts on a current pyloric motor pattern. Moreover, it might render the motoneurones of the pyloric pattern generator more sensitive to inputs from a command oscillator, and contribute to switching on the pyloric motor pattern.