Outbreak of Pseudomonas aeruginosa producing VIM carbapenemase in an intensive care unit and its termination by implementation of waterless patient care.

BACKGROUND: Long-term outbreaks of multidrug-resistant Gram-negative bacilli related to hospital-building water systems have been described. However, successful mitigation strategies have rarely been reported. In particular, environmental disinfection or replacement of contaminated equipment usually failed to eradicate environmental sources of Pseudomonas aeruginosa. METHODS: We report the investigation and termination of an outbreak of P. aeruginosa producing VIM carbapenemase (PA-VIM) in the adult intensive care unit (ICU) of a Swiss tertiary care hospital with active case finding, environmental sampling and whole genome sequencing (WGS) of patient and environmental strains. We also describe the implemented control strategies and their effectiveness on eradication of the environmental reservoir. RESULTS: Between April 2018 and September 2020, 21 patients became either infected or colonized with a PA-VIM strain. For 16 of them, an acquisition in the ICU was suspected. Among 131 environmental samples collected in the ICU, 13 grew PA-VIM in sink traps and drains. WGS confirmed the epidemiological link between clinical and environmental strains and the monoclonal pattern of the outbreak. After removing sinks from patient rooms and implementation of waterless patient care, no new acquisition was detected in the ICU within 8 months after the intervention. DISCUSSION: Implementation of waterless patient care with removal of the sinks in patient rooms was successful for termination of a PA-VIM ICU outbreak linked to multiple environmental water sources. WGS provides highly discriminatory accuracy to investigate environment-related outbreaks.

Randomized, placebo-controlled, double-blind clinical trial to evaluate the efficacy of polyhexanide for topical decolonization of MRSA carriers.

OBJECTIVES: The objective of this study was to evaluate the efficacy of polyhexanide (Prontoderm(®)) in eliminating MRSA carriage. METHODS: In a 1900 bed teaching hospital, MRSA-colonized patients were randomized into a double-blind, placebo-controlled superiority trial between January 2011 and July 2014. Patients were treated with either polyhexanide or placebo applied to the anterior nares (thrice daily) and skin (once daily) for 10 days. The primary outcome was MRSA decolonization at day 28 (D28) after the end of treatment assessed by ITT responder and PP analyses (microbiological follow-up ± 7 days and topical treatment ≥ 5 days). Secondary outcomes included safety, emergence of resistance and MRSA genotype changes. Registered trial number ISRCTN02288276. RESULTS: Of 2590 patients screened, 146 (polyhexanide group, 71; placebo group, 75) were included. ITT analysis showed that 24/71 (33.8%) patients in the polyhexanide group versus 22/75 (29.3%) in the placebo group were MRSA-free at D28 (risk difference, 4.5%; 95% CI, -10.6% to 19.5%; P = 0.56). PP analysis confirmed the results with 19/53 (35.8%) decolonized polyhexanide-treated patients versus 17/56 (30.4%) in the placebo arm (risk difference, 5.5%; 95% CI, -12.2% to 23%; P = 0.54). Nine serious adverse events occurred in the polyhexanide group versus 12 in the placebo group; none was attributable to study medication. Emergence of polyhexanide resistance or cross-resistance between polyhexanide and chlorhexidine was not observed. No case of exogenous recolonization by a genotypically different MRSA strain was documented. CONCLUSIONS: This study suggests that under real-life conditions, a single polyhexanide decolonization course is not effective in eradicating MRSA carriage.

Ampicillin-resistant Haemophilus influenzae isolates in Geneva: serotype, antimicrobial susceptibility, and ß-lactam resistance mechanisms.

The purpose of this study was to analyze the molecular mechanisms of ampicillin-resistant Haemophilus influenzae isolated in Geneva, Switzerland. We investigated the association between specific patterns of amino acid substitutions in penicillin-binding protein 3 (with or without ß-lactamase production) and ß-lactam susceptibility. Another main focus for this study was to compare the accuracy of disk diffusion and Etest methods to detect resistance to ampicillin and amoxicillin/clavulanic acid. The antibiotic susceptibility to ß-lactam antibiotics of 124 H. influenzae isolates was determined by disk diffusion and Etest methods, and interpreted by European Committee on Antimicrobial Susceptibility Testing (EUCAST) and Clinical and Laboratory Standards Institute (CLSI) breakpoints. Alterations in PBP3 were investigated by sequencing the ftsI gene. Of the 124 clinical isolates analyzed, ampicillin resistance was found in 36% (45 out of 124). The rate of resistance to amoxicillin/clavulanic acid was 9% and 0.8%, using EUCAST and CLSI breakpoints respectively. For the 78 ß-lactamase negative ampicillin-susceptible (BLNAS) isolates for which the Etest method indicated a high degree of susceptibility (MIC ≤ 1 mg/L), the disk diffusion method revealed resistance to ampicillin and amoxicillin/clavulanic acid in 33 cases (42%). Most common amino acid substitutions were Asn526Lys and Val547Ile, followed by Asp569Ser, Ala502Val, Asp350Asn, Met377Ile, Ile449Val, and Arg517His. The patterns observed were classified into six groups (IIa, IIb, IIc, IId, III-like, and miscellaneous). Continued characterization of both invasive and respiratory H. influenzae isolates is necessary in order to observe changes in the microbiology and epidemiology of this pathogen that could lead to clinical failure when treated by empirical antibiotic therapy.

[Diagnosis of bacterial gastroenteritis]. / Diagnostic de la gastroentérite bactérienne.

The etiologic agents of acute gastroenteritis are diverse. The diagnosis of bacterial pathogens is particularly challenging given the large amount of vastly diverse indigenous gastrointestinal organisms present in stool. Multiple methods must be used by the clinical microbiology laboratories to diagnose the cause of acute gastroenteritis, including bacterial cultures, ELISA, and microscopy. Due to the limitations of conventional methods, there is still room for improvement in the detection of pathogens by using the molecular methods. This paper discusses these different diagnostic approaches and limitations.

[ESBL and carbapenemases in Enterobacteriaceae]. / Bêtalactamases à spectre étendu et carbapénémases chez les Enterobacteriaceae.

Carbapenemase-producing enterobacteria (CPE) spread all over the world during the last years, causing serious infections with increasing frequency. Very few new drugs active against CPE are expected to be clinically available. Studies summarized in this review show that there is yet room to improve our therapeutic approaches, in the treatment of infections due to CPE.

Targeted mupirocin-based decolonization for Staphylococcus aureus carriers and the subsequent risk of mupirocin resistance in haemodialysis patients - a longitudinal study over 20 years.

Mupirocin-based decolonization of Staphylococcus aureus carriers undergoing haemodialysis is not widely implemented due to concerns of mupirocin resistance. In our haemodialysis unit, a strategy combining universal S. aureus screening with targeted mupirocin-based decolonization was introduced two decades ago. In this study of haemodialysis patients, mupirocin resistance was assessed in blood and colonizing S. aureus isolates during two periods. Mupirocin resistance in S. aureus was infrequent in both blood and colonizing isolates. Furthermore, in the years 2003-2021, a decreasing trend in the annual rate of S. aureus bloodstream infections was observed. Targeted mupirocin-based decolonization of S. aureus carriers undergoing haemodialysis is a sustainable measure for preventing healthcare-associated infections.

Comparison of the Copan WASPLab incorporating the BioRad expert system against the SIRscan 2000 automatic for routine antimicrobial disc diffusion susceptibility testing.

OBJECTIVES: This study investigated the agreement at the categorical level between the Copan WASPLab incorporating the BioRad expert system against the SIRscan 2000 automatic for antimicrobial disc diffusion susceptibility testing. METHODS: The 338 clinical strains (67 Pseudomonas aeruginosa, 19 methicillin-resistant Staphylococcus aureus, 75 methicillin-sensitive S. aureus and 177 Enterobacterales isolates) analysed in this study were non-duplicate isolates obtained from consecutive clinical samples referred to the clinical bacteriology laboratory at Geneva University Hospitals between June and August 2019. For the WASPLab the inoculum suspension was prepared in strict accordance with the manufacturer's instruction (Copan WASP srl, Brescia, Italy) by adding 2 mL of the 0.5 McFarland primary suspension used for the SIRscan analysis into a sterile tube filled with 4 mL of sterile saline (1:3 dilution). The inoculum (2 × 30 µL loop/spreader) was spread over the entire surface of Mueller-Hinton agar plates according to the AST streaking pattern defined by Copan. The antibiotic discs were dispensed by the WASP and inoculated media were loaded on conveyors for transfer to the automatic incubators. The plates were incubated for 16 h, and several digital images were acquired. Inhibition zone diameters were automatically read by the WASPLab and were adjusted manually whenever necessary. For the SIRscan 2000 automatic, the antimicrobial disc diffusion susceptibility testing was performed according to the EUCAST guidelines. The gradient strip method was used to resolve discrepancies. RESULTS: The overall categorical agreement between the compared methods reached 99.1% (797/804; 95% CI 98.2%-99.6%), 99.5% (1029/1034; 95% CI 98.9%-99.8%), and 98.8% (2798/2832; 95% CI 98.3%-99.1%) for P. aeruginosa, S. aureus and the Enterobacterales, respectively. CONCLUSIONS: WASPLab incorporating the BioRad expert system provides a fully automated solution for antimicrobial disc diffusion susceptibility testing with equal or better accuracy than other available phenotypic methods.

Copan WASPLab automation significantly reduces incubation times and allows earlier culture readings.

OBJECTIVE: The aim was to evaluate whether laboratory automation (inoculation and automated incubation combined with timely defined high-resolution digital imaging) may help reduce the time required to obtain reliable culture analysis results. METHODS: We compared the results obtained by WASPLab automation against WASP-based automated inoculation coupled to conventional incubation and manual diagnostic on 1294 clinical samples (483 for the derivation set and 811 for the independent validation set) that included urine, genital tract and non-sterile site specimens, as well as ESwabs for screening of methicillin-resistant Staphylococcus aureus (MRSA), methicillin-sensitive Staphylococcus aureus (MSSA), extended-spectrum beta-lactamases (ESBLs) and carbapenemase-producing Enterobacteriaceae (CPE). We used sequential routine specimens referred to the bacteriology laboratory at Geneva University Hospitals between October 2018 and March 2019. RESULTS: The detection sensitivity of MRSA and MSSA at 18 hr on WASPLab was 100% (95% confidence interval [CI], 94.48-100.00%). The detection sensitivity of ESBL and CPE at 16 hr on WASPLab was 100% (95% confidence interval [CI], 94.87% to 100.00%). For urine specimens, the similarity was 79% (295/375) between 18 hr and 24 hr of incubation on WASPLab. For genital tract and non-sterile site specimens, the similarity between 16 hr and 28 hr of incubation on WASPLab were 26% (72/281) and 77% (123/159) respectively. Thus, 28 hr was defined as the final incubation time on WASPLab for genital tract and non-sterile site specimens. CONCLUSIONS: The results of this study show that WASPLab automation enables a reduction of the culture reading time for all specimens tested without affecting performances. Implementing the established and duly validated incubation times will allow appropriate laboratory workflows for improved efficiency to be built.

A molecular based diagnosis of positive blood culture in the context of viral haemorrhagic fever: proof of concept.

OBJECTIVES: The aim of this study was to evaluate the possibility of using a PCR-based panel to identify bacterial and fungal bloodstream infections in the setting of suspected or confirmed viral haemorrhagic fever. METHODS: The accuracy of the FilmArray® Blood Culture Identification Panel (BCID) assay was assessed to identify the common bacterial and fungal pathogens associated with bloodstream infections after positive blood culture inactivation using a guanidinium thiocyanate containing buffer lysis that is commonly used for viral haemorrhagic fever molecular diagnostics. RESULTS: The FilmArray® BCID panel assay detected 95% (19/20) of the pathogens analysed in this study by using both protocols with and without inactivation. Absolute consistency (100%) was observed in all isolates with phenotypes compatible with the presence of the antibiotic resistance genes mecA, vanA, vanB and blaKPC. CONCLUSIONS: The FilmArray® BCID panel assay coupled to inactivation using a guanidinium thiocyanate containing buffer lysis represents a convenient, sensitive and specific diagnostic tool to detect some of the most pathogens associated with bloodstream infections in the context of a suspected or confirmed viral haemorrhagic fever.

Imipenem heteroresistance in nontypeable Haemophilus influenzae is linked to a combination of altered PBP3, slow drug influx and direct efflux regulation.

OBJECTIVE: To investigate the potential roles of PBPs, efflux pumps and slow drug influx for imipenem heteroresistance in nontypeable Haemophilus influenzae (NTHi). METHODS: Fifty-nine NTHi clinical isolates examined in this study were collected at Geneva University Hospitals between 2009 and 2014. Alterations in PBPs were investigated by gene sequencing. To evaluate the affinities of the PBPs to imipenem, steady-state concentration-response experiments were carried out using imipenem in a competition assay with Bocillin-FL. The effect of the carbonyl cyanide m-chlorophenylhydrazone (CCCP) on imipenem susceptibility was assessed using broth dilution and viable cell counting. Using whole-genome sequencing, we explored the potential roles of outer membrane protein P2 (OmpP2), LytM proteins and the dcw gene cluster in imipenem heteroresistance. RESULTS: All 46 imipenem-heteroresistant isolates (IMIhR) harboured amino acid substitutions in the ftsI gene, which encodes PBP3, corresponding to 25 different mutation patterns that varied from the ftsI gene mutation patterns found in imipenem-susceptible isolates. Among all PBPs, the highest affinity to imipenem was documented for PBP3 (IC50, 0.004 µg/mL). Different amino acid substitutions and insertions were noted in OmpP2, suggesting a relationship with imipenem heteroresistance. The IMIhR isolates were affected by CCCP differently and displayed a higher percentage of killing by imipenem in CCCP-treated cells at concentrations ranging between 0.5 and 8 µg/mL. CONCLUSIONS: The present study provides robust evidence indicating that in combination with the altered PBP3, the slowed drug influx and its enhanced efflux due to the loss of regulation led to the development of imipenem heteroresistance in NTHi.

Clonal or not clonal? Investigating hospital outbreaks of KPC-producing Klebsiella pneumoniae with whole-genome sequencing.

OBJECTIVES: Whole-genome sequencing (WGS) is a promising tool for identifying transmission pathways in outbreaks caused by multidrug-resistant bacteria. However, it is uncertain how the data produced by WGS can be best integrated into epidemiologic investigations. METHODS: We tested various genomic analyses to identify clonal groups in two distinct outbreaks of Klebsiella pneumoniae carbapenemase-producing K. pneumoniae that occurred in Switzerland in 2013 and 2015. In blinded fashion, we sequenced 12 strains involved in the two outbreaks, respectively, and six that were epidemiologically unrelated. We analysed genomic commonalities from conserved genes to plasmid-borne antibiotic resistance genes (ARGs) and contrasted these results with available epidemiologic evidence. RESULTS: Using WGS, blinded analysts correctly identified the two clusters of strains from the two outbreaks. Nonetheless, the 2015 index strain was found to be slightly different (1-3 single nucleotide variants) from the strains recovered from secondary cases, likely because prior long-term carriage (3 years) by the index patient allowed for genetic mutations over time. Also, we observed occasional loss of ARG-bearing plasmidic fragments in outbreak-causing strains. CONCLUSIONS: Retrospective WGS analysis was successful in identifying clonal groups in both outbreaks. Still, data should be analysed with caution in cases of previous long-term carriage of the studied bacteria.

Community-acquired methicillin-resistant Staphylococcus aureus in Switzerland : first surveillance report.

Methicillin resistant Staphylococcus Aureus (MRSA) infection is an emerging community pathogen. Community-acquired MRSA (CA-MRSA) has been associated with virulent strains producing Panton-Valentine leukocidin (PVL) and a variety of other exotoxins. In Geneva, PVL-producing CA-MRSA was first reported in 2002 and a surveillance system based on voluntary reporting was set up.

Community-acquired methicillin-resistant Staphylococcus aureus in Switzerland: first surveillance report.

Methicillin-resistant Staphylococcus aureus (MRSA) is an emerging community pathogen. Community-acquired MRSA (CA-MRSA) has been associated with virulent strains producing Panton-Valentine leukocidin (PVL) and a variety of other exotoxins. In Geneva, PVL-producing CA-MRSA was first reported in 2002 and a surveillance system based on voluntary reporting was set up. Each MRSA-positive culture result with an antibiotic resistance profile different from the endemic strain prevailing in the Geneva healthcare setting diagnosed in a patient without a history of hospital admission in the previous 12 months was notified to the local health department. A questionnaire was completed by the attending physician with demographic, clinical and exposure information. From January 2002 until December 2004, data on 58 cases were reported, including 26 cases grouped in 13 distinct transmission clusters. Most were family related and for two of them, colonisation persisted over a 12 month period despite treatment. Thirty three patients (57%) were male. Median age was 32 years, 22% being younger than 10 years. Forty one cases (71%) were infected and 17 (29%) colonised. Symptomatic skin lesions such as furunculosis, impetigo or abscess were present in 40 (97%) of the 41 infected cases. Most cases had no underlying disease. Thirty eight cases (65%) had travelled abroad. Forty (69%) of 58 isolates carried the PVL toxin. CA-MRSA infections in Geneva appear to be an emerging problem in the canton. Surveillance should continue and should possibly be extended to other parts of the country to better describe transmission patterns and the spread of this pathogen. Prevention and control of CA-MRSA infections represent a challenge for the future, requiring contact tracing, education and treatment of infected and colonised contacts.

Rapid molecular determination of methicillin resistance in staphylococcal bacteraemia improves early targeted antibiotic prescribing: a randomized clinical trial.

Empiric therapy of methicillin-susceptible Staphylococcus aureus (MSSA) infections with vancomycin is associated with poorer outcome than targeted therapy with ß-lactams. Our objective was to evaluate whether rapid determination of methicillin resistance shortens the time from Gram stain to targeted antimicrobial therapy in staphylococcal bacteraemia, thereby reducing vancomycin overuse. This was a single-centre open parallel RCT. Gram-positive cocci in clusters in positive blood culture underwent real-time PCR for rapid species and methicillin resistance determination parallel to conventional microbiology. Patients were randomized 1:1 so that clinicians would be informed of PCR results (intervention group) or not (control group). Eighty-nine patients (intervention 48, control 41) were analysed. MRSA was identified in seven patients, MSSA in 46, and CoNS in 36. PCR results were highly concordant (87/89) with standard microbiology. Median time (hours) from Gram stain to transmission of methicillin-susceptibility was 3.9 (2.8-4.3) vs. 25.4 (24.4-26-7) in intervention vs. control groups (p <0.001). Median time (hours) from Gram stain to targeted treatment was similar for 'all staphylococci' [6 (3.8-10) vs. 8 (1-36) p 0.13] but shorter in the intervention group when considering S. aureus only [5 (3-7) vs. 25.5 (3.8-54) p <0.001]. When standard susceptibility testing was complete, 41/48 (85.4%) patients in the intervention group were already receiving targeted therapy compared with 23/41 (56.1%) in the control group (p 0.004). There was no significant effect on clinical outcomes. Rapid determination of methicillin resistance in staphylococcal bacteraemia is accurate and reduces significantly the time to targeted antibiotic therapy in the subgroup of S. aureus, thereby avoiding unnecessary exposure to vancomycin.

Reversible inhibition of acetylcholinesterase by eseroline, an opioid agonist structurally related to physostigmine (eserine) and morphine.

The action of eseroline--(3aS,8aR)-1,2,3,3a,8,8a-hexahydro-1,3a,8-trimethylpyrrolo[2,3-b]indo l-5-ol--salicylate was tested on preparations of ChE from different sources and on the longitudinal muscle of guinea-pig ileum. While eseroline is eseroline is extremely weak-acting on horse serum BuChE (Ki = 208 +/- 42 microM), it is a rather strong competitive inhibitor of AChE's, its Ki being 0.15 +/- 0.08 microM, 0.22 +/- 0.10 microM and 0.61 +/- 0.12 microM in electric eel, human RBC and rat brain, respectively. Eseroline inhibitory action in AChE in independent of the duration of pre-incubation and appears fully developed in less than 15 sec. This action is also rapidly reversible; after pre-incubation followed by dilution, maximum enzymic activity is regained within 15 sec. The electrically-evoked contractions of the longitudinal strip were inhibited by concentrations of eseroline in the range 0.2-15 microM, while they were increased by concentrations over 20 microM. In the same preparation, without electrical stimulation, but in the presence of naloxone, eseroline induced contractions at concentrations higher than 5 microM. This effect was antagonized by atropine. The inhibitory activity of eseroline parallels, as regards selectivity, potency and kinetics, that of the phenolic anticurare agent edrophonium, while it differs markedly from that of physostigmine.

Breathing pattern in sarcoidosis and idiopathic pulmonary fibrosis.

We measured the volume and time components of the breathing cycle together with the P.1 in 16 sarcoidosis and 9 IPF patients. Patients in the IPF group were older, and their lung functions revealed a more restrictive pattern with small TLC, VC, and higher elastance values. We observed that in the two groups of patients there was a significant correlation between VT, f, and P.1 and lung elastance, and that the stiffer IPF patients had higher drive parameters. Hence, the main factor affecting the breathing pattern in both groups is the influence of the elastic load on the control of breathing. A significant relationship was also found between %VD/VT and elastance. We then compared the f values of the 16 sarcoidosis and 9 IPF patients with those of elastically loaded normal subjects. At equivalent levels of elastance, even though the patients and the normals showed qualitatively similar changes, the patients tended to have higher frequencies for the equivalent elastic load. In conclusion, ventilatory and drive parameters in interstitial lung diseases increased as a function of the elastic load, this load being greater in IPF than in sarcoidosis. Inflammation may stimulate the vagal receptors to increase the frequency of breathing, but this requires further study.

Eseroline: a new antinociceptive agent derived from physostigmine with opiate receptor agonist properties. Experimental in vivo and in vitro studies on cats and rodents.

We report that eseroline, until now thought devoid of any biological action, is a potent antinociceptive agent. Its antinociceptive action is stronger than that of morphine in all tests studied and, though shorter lasting than that of the latter, has a latency of only a few minutes by subcutaneous route. Eseroline, like morphine and enkephalins, inhibits the electrically evoked twitches of the mouse vas deferens and of the guinea-pig ileum. Eseroline, moreover, releases 5-hydroxytryptamine from cat brain cortex in way similar to that of morphine and physostigmine.

Complexes between pilocarpine and cobalt(II), nickel(II), copper(II), and zinc(II) ions.

The properties of pilocarpine as a ligand toward the halides of cobalt(II), nickel(II), copper(II), and zinc(II) were investigated. Pilocarpine behaved as a monodentate ligand, giving rise to compounds with the general formula methyl(pilocarpine)2X2. The coordinating geometry at the metal ion was pseudotetrahedral in every case. PMR studies showed that the pyridine-type nitrogen of the imidazole ring of pilocarpine was the donor atom of the ligand.

Genotoxicity assay of oil dispersants in bacteria (mutation, differential lethality, SOS DNA-repair) and yeast (mitotic crossing-over).

5 oil dispersants and a sample of paraffin were devoid of mutagenic activity in the Ames reversion test, with and without S9 mix, using 7 his- S. typhimurium strains (TA1535, TA1537, TA1538, TA97, TA98, TA100, TA102). However, 3 dispersants produced direct DNA damage in E. coli WP2, which was not repairable in repair-deficient strains (WP2uvrA, CM871, TM1080), as shown by two different DNA-repair test procedures. The uvrA excision-repair system was in all cases the most important mechanism involved in repairing the DNA damage produced by oil dispersants, while the combination of uvrA with other genetic defects (polA, recA, lexA) decreased the efficiency of the system. The observed genotoxic effects were considerably lowered in the presence of S9 mix containing liver S9 fractions from Aroclor-treated rats. The sample of oil dispersant yielding the most pronounced DNA damage in repair-deficient E. coli failed to induce gene sfiA in E. coli (strain PQ37), using the SOS chromotest, or mitotic crossing-over in Saccharomyces cerevisiae (strain D5). The direct toxicity of the oil dispersant to both bacterial and yeast cells was markedly decreased in the presence of rat-liver preparations. These two short-term tests were effective in detecting the genotoxicity of both direct-acting compounds (such as 4-nitroquinoline N-oxide and methyl methanesulfonate) and procarcinogens (such as cyclophosphamide, 2-aminoanthracene and 2-aminofluorene). Moreover, the SOS chromotest was successfully applied to discriminate the activity of chromium compounds as related to their valence (i.e. Cr(VI) genotoxic and Cr(III) inactive). Combination of oil dispersants with Cr(VI) compounds did not affect the direct mutagenicity to S. typhimurium (TA102) of a soluble salt (sodium dichromate) nor did it result in any release of a water-soluble salt (lead chromate), as also confirmed by analytical methods. On the other hand, exposure to sunlight tended to decrease, to a slow rate, the direct genotoxicity of an oil dispersant in the bacterial DNA-repair test.

In-vitro and in-vivo protection of acetylcholinesterase by eseroline against inactivation by diisopropyl fluorophosphate and carbamates.

The protective action of eseroline--(3aS,8aR)-1,2,3,3a,8,8a-hexahydro-1,3 a, 8-trimethyl-pyrrolo[2,3-b]indol-5-ol--salicylate against (DFP) diisopropyl fluorophosphate and carbamate poisoning of cholinesterases (ChEs) has been examined in-vitro with human erythrocytes and purified preparations of electric eel acetylcholinesterase (AChE) and of horse serum butyrylcholinesterase (BuChE), and in-vivo using mice. Eseroline afforded 50% protection (ED 50) of erythrocyte AChE against inactivation by 1 microM DFP, physostigmine or neostigmine, at concentrations of 4.3, 22 and 23.5 microM, respectively, while for eel AChE protection against 10 and 30 microM DFP, 0.3 and 1 microM physostigmine and 1 microM neostigmine the eseroline ED 50 values were 0.3, 0.4, 0.7, 1.9 and 5.6 microM, respectively. On the other hand, up to 0.3 mM eseroline did not appreciably affect the inhibitory action of the same drugs on horse serum BuChE. Eseroline concentrations in the range 0.1-1 mM were able to reactivate 20-42% of erythrocyte AChE previously inhibited by 100 microM physostigmine, but failed to reactivate the DFP (10 microM)-pretreated enzyme to any extent. Finally, eseroline salicylate injected into mice (10 mg kg-1 s.c.) protected an average of 82 and 26% of the animals against lethal doses of DFP (7 mg kg-1 s.c.) and physostigmine sulphate (1 mg kg-1 i.p.) respectively, which were administered 15 min later. These results indicate that the protective activity of eseroline correlates well with its own anti-ChE profile, and that the effectiveness of the protection depends largely on the rate of AChE inhibition by the agents used to inactivate the enzyme.