Rafael Gómez-Lus

Bacteriocin Production in Vancomycin-Resistant and Vancomycin-Susceptible Enterococcus Isolates of Different Origins

ABSTRACT Bacteriocin production was determined for 218Enterococcus isolates (Enterococcus faecalis[93] and E. faecium[125]) obtained from different origins (human clinical samples [87], human fecal samples [78], sewage [28], and chicken samples [25]) and showing different vancomycin susceptibility patterns (vancomycin resistant, all of them vanA positive [56], and vancomycin susceptible [162]). All enterococcal isolates were randomly selected except for the vancomycin-resistant ones. A total of 33 isolates of eight different bacterial genera were used as indicators for bacteriocin production. Forty-seven percent of the analyzed enterococcal isolates were bacteriocin producers (80.6% of E. faecalis and 21.6% ofE. faecium isolates). The percentage of bacteriocin producers was higher among human clinical isolates (63.2%, 81.8% of vancomycin-resistant isolates and 60.5% of vancomycin-susceptible ones) than among isolates from the other origins (28 to 39.3%). Only one out of the 15 vancomycin-resistant isolates from human fecal samples was a bacteriocin producer, while 44.4% of fecal vancomycin-susceptible isolates were. The bacteriocin produced by the vanA-containing E. faecium strain RC714, named bacteriocin RC714, was further characterized. This bacteriocin activity was cotransferred together with thevanA genetic determinant to E. faecalis strain JH2-2. Bacteriocin RC714 was purified to homogeneity and its primary structure was determined by amino acid sequencing, showing an identity of 88% and a similarity of 92% with the previously described bacteriocin 31 from E. faecalis YI717. The presence of five different amino acids in bacteriocin RC714 suggest that this could be a new bacteriocin. The results obtained suggest that the epidemiology of vancomycin resistance may be influenced by different factors, including bacteriocin production.

Molecular basis of resistance to macrolides and other antibiotics in commensal viridans group streptococci and Gemella spp. and transfer of resistance genes to Streptococcus pneumoniae.

ABSTRACT We assessed the mechanisms of resistance to macrolide-lincosamide-streptogramin B (MLSB) antibiotics and related antibiotics in erythromycin-resistant viridans group streptococci (n = 164) and Gemella spp. (n = 28). The macrolide resistance phenotype was predominant (59.38%); all isolates with this phenotype carried the mef(A) or mef(E) gene, with mef(E) being predominant (95.36%). The erm(B) gene was always detected in strains with constitutive and inducible MLSB resistance and was combined with the mef(A/E) gene in 47.44% of isolates. None of the isolates carried the erm(A) subclass erm(TR), erm(A), or erm(C) genes. The mel gene was detected in all but four strains carrying the mef(A/E) gene. The tet(M) gene was found in 86.90% of tetracycline-resistant isolates and was strongly associated with the presence of the erm(B) gene. The catpC194 gene was detected in seven chloramphenicol-resistant Streptococcus mitis isolates, and the aph(3′)-III gene was detected in four viridans group streptococcal isolates with high-level kanamycin resistance. The intTn gene was found in all isolates with the erm(B), tet(M), aph(3′)-III, and catpC194 gene. The mef(E) and mel genes were successfully transferred from both groups of bacteria to Streptococcus pneumoniae R6 by transformation. Viridans group streptococci and Gemella spp. seem to be important reservoirs of resistance genes.

Respiratory cryptosporidiosis: Case series and review of the literature

SummaryFive case of intestinal cryptosporidiosis with pulmonary involvement in patients with AIDS are reported. The diagnosis was based on the recognition of acid-fast oocysts in sputum or aspirated bronchial material and stool specimens. Coughing and excess secretions were present in all cases. Four patients had other associated pulmonary pathogens: twoMycobacterium tuberculosis, oneMycobacterium fortuitum and one Cytomegalovirus +Pneumocystis carinii; all of them had a previous (three cases) or simultaneous (one case) diagnosis of intestinal cryptosporidiosis, presenting with diarrhoea and vomiting. In the fifth patientCryptosporidium was the only pulmonary pathogen found in a bronchial aspirate, and the onset of diarrhoea was 1 month after respiratory detection. Fifty-seven cases of respiratory cryptosporidiosis have been reported since 1980. In 17 of them, no other pathogen was found. Diarrhoea was present in 77% of the patients, cough in 77%, dyspnea in 58%, expectoration in 54%, fever in 45%, thoracic pain in 33%.ZusammenfassungWir berichten über fünf Fälle von intestinaler Kryptosporidiose mit pulmonaler Beteiligung bei AIDS Patienten. Die Diagnose wurde durch den Nachweis von säurefesten Oozysten im Sputum oder aspiriertem Bronchialsekret und im Stuhl gestellt. Bei allen Patienten bestand Husten und Auswurf. Bei vier Patienten fand sich gleichzeitig eine Assoziation mit anderen Infektionen, in zwei Fällen durchMycobacterium tuberculosis, in einem Fall durchMycobacterium fortuitum und in einem Fall durch Cytomegalovirus plusPneumocystis carinii. Bei allen Patienten war eine vorbestehende (drei Fälle) oder gleichzeitige intestinale Kryptosporidiose vorhanden, die mit Durchfall und Erbrechen einherging. Bei dem fünften Patienten war im Bronchialsekret ausschließlichCryptosporidium nachzuweisen, die Diarrhoe setzte erst einen Monat nach Entdeckung des Erregers im Respirationstrakt ein. Seit 1980 wurden 57 Fälle von respiratorischer Kryptosporidiose publiziert. In 17 Fällen fand sich kein anderer Erreger. In je 77% der Fälle waren Diarrhoe und Husten, in 58% Atemnot, in 54% Auswurf, 45% Fieber und in 33% Schmerzen aufgetreten.

Western blot applied to the diagnosis and post-treatment monitoring of human hydatidosis.

The serologic diagnosis of hydatidosis (caused by Echinococcus granulosus) can be made by different techniques, although the lack of standardization of the antigens affects the sensitivity, specificity and concordance among the different tests. We have applied the Western-Blot (WB) technique, associated with a purified antigen from sheep hydatid fluid, at 60 samples of serum from 14 patients suffering echinococcosis in different bodily locations, monitored for 3 years. The WB test enabled the detection of antibodies in the pre-surgical samples for proteins of 12-14, 16, 20, 24-26, 34, 39 and 42 kDa in molecular weight in 15-96% of the patients. The combination involving 2 of the 3 proteins of 20, 39 and 42 kDa has made it possible to diagnose 100% of the cases. The antibodies specific to proteins 39 and 42 kDa disappeared in less than one year in the patients cured after surgery, while in patients with persistent or recurrent parasitism the bands present before surgery persisted or other new ones appeared. The WB with purified antigens proved to be highly useful in the diagnosis and post-surgical monitoring of hydatidosis patients. The antigen used is proposed as a standard antigen for the diagnosis and follow-up of pre- and postsurgical hydatidosis.

Evaluation of Bacterial Resistance to Essential Oils and Antibiotics After Exposure to Oregano and Cinnamon Essential Oils

Essential oils (EOs) are excellent antimicrobial agents sometimes used in active food packaging. This work studies the susceptibility of 48 clinical isolates and 12 reference strains of Gram-negative bacilli to oregano essential oil, cinnamon essential oil, and combinations of both. Furthermore, the tendency of the clinical isolates to develop resistance to these EOs and to different antibiotics after sequential oregano or cinnamon exposure was studied. For this purpose, antibiotic susceptibility (through disk diffusion assays and minimum inhibitory concentration [MIC] determination) and oregano and cinnamon susceptibility (through MIC and minimum bactericidal concentration [MBC] determination) were compared after 50 passages in the presence or absence of subinhibitory concentrations of oregano and cinnamon essential oils. The results showed that all strains were susceptible to both EOs and their combination independently of the antibiotic resistance profile. In addition, neither synergistic nor antagonistic effects were observed between oregano and cinnamon essential oils at the concentrations tested. After the sequential exposure to both EOs, only Serratia marcescens, Morganella morganii, and Proteus mirabilis treated with oregano changed their antibiotic resistance profile and/or increased their resistance to this EO. However, the changes in antibiotic and oregano resistance were not related.

Aminoglycoside acetyltransferase 3-IV (aacC4) and hygromycin B 4-I phosphotransferase (hphB) in bacteria isolated from human and animal sources.

Members of the family Enterobacteriaceae harboring an enzyme of the aminoglycoside acetyltransferase 3 class (AAC-3-IV) (apramycin and gentamicin resistance) and hygromycin B phosphotransferase 4 (HPH-4-I) (hygromycin B resistance) have been isolated from human clinical sources in Europe. A cluster of genes containing IS140, aacC4, and hphB was found in these strains. We demonstrate by Southern hybridization that this cluster is identical to the operon found in animals that also contains insertion sequences belonging to the ISO family. This provides another example of presumptive transfer of antibiotic resistance genes between bacteria of animal and human origin. Images

Evaluation of an Immunochromatographic Dip-Strip Test for the Detection of Cryptosporidium Oocysts in Stool Specimens

Abstract.The study presented here examined the efficacy of a commercially available qualitative immunochromatographic assay for detecting Cryptosporidium oocysts in stool samples. A total of 75 samples were tested, including 50 positive for Cryptosporidium spp. by acid-fast stain, 20 positive for other parasites (Blastocystis hominis, Endolimax nana, Entamoeba coli, Giardia lamblia, Ascaris lumbricoides, Strongyloides stercoralis and Trichuris trichiura), and five negative samples. The observed sensitivity was 98%, while specificity was 100%; the detection threshold was near 1,000 oocysts/ml. Correctly diagnosed positive samples included Cryptosporidium parvum genotypes 1 and 2, whereas the single false-negative sample corresponded to a Cryptosporidium meleagridis infection.

EPIDEMIOLOGICAL MARKERS OF ACINETOBACTER BAUMANNII CLINICAL ISOLATES FROM A SPINAL CORD INJURY UNIT

During a period of 28 months, 114 isolates of Acinetobacter baumannii obtained from urine samples of 57 patients, were recovered in a Spinal Cord Unit; an unusual increase in the number of A. baumannii isolates was observed between February 1991 and January 1992. Six different typing methods [biotyping, antimicrobial susceptibility, whole cell and cell-envelope protein analysis, plasmid analysis and chromosomal DNA analysis by pulsed-field gel electrophoresis (PFGE)] were used to study the isolates to establish any potential relationships among them. Chromosomal DNA analysis by digestion with ApaI and separation of the fragments by PFGE was the most powerful tool to determine the relatedness of isolates. The results suggest that the isolates from 1991 and 1992 may have originated from strains present in 1990 that subsequently acquired resistance to amikacin and tobramycin during the epidemic.

Comparative in vitro bacteriostatic and bactericidal activity of trovafloxacin, levofloxacin and moxifloxacin against clinical and environmental isolates of Legionella spp.

The susceptibility of 140 Legionella spp isolates (106 clinical and 34 environmental isolates) to trovafloxacin (TRFX), levofloxacin (LEVX), moxifloxacin (MOFX), ciprofloxacin (CIPX), ofloxacin (OFLX), erythromycin (ERY), azithromycin (AZI) and rifampicin (RIF) was studied using a standard microdilution method and buffered yeast extract broth (BYE) supplemented with 0.1% alpha-ketoglutarate. The post-antibiotic effects (PAEs) of the study drugs against 10 clinical isolates of Legionella pneumophila sg.1 were compared. The MIC inhibiting 90% of strains tested on BYEalpha broth were 0.008, 0.016, 0.016, 0.06, 0.125, 0.5, 0.5, and 0.004 mg/l for TRFX, LEVX, MOXX, CIP, OFLX, ERY, AZI, and RIF, respectively. The MBC/MIC ratios ranged from one to eight depending on the antibiotic tested: TRFX [1x-2 x MIC], LEVX, MOFX, CIPX and OFLX [1x-4 x MIC], RIF [2x-4 x MIC], ERY and AZI [2x-8 x MIC]. TRFX, RIF, LEVX, MOFX, CIPX, OFLX, ERY and AZI showed similar activity against Legionella species other than L. pneumophila. One-hour exposures to the study antimicrobial agents at a concentration of 4 x MIC resulted in PAEs as follows (average in hours): TRFX: 2.68 h; RIF: 2.63 h; CIPX: 2.62 h; MOFX: 2.56 h; LEVX: 2.41 h; OFLX: 2.25 h; AZI: 1.65 h; and ERY: 1.54 h. In conclusion, our in vitro data confirm that trovafloxacin, levofloxacin, moxifloxacin and rifampicin have excellent bacteriostatic and bactericidal activity against Legionella spp and show significant post-antibiotic effect.

Epidemiological study of resistance to nalidixic acid and other antibiotics in clinical Yersinia enterocolitica O:3 isolates.

ABSTRACT Forty-six Yersinia enterocolitica O:3 clinical isolates resistant to nalidixic acid were studied. The use of molecular typing techniques, other indicators of resistance patterns, the plasmid profile, and the presence of genes that encode aminoglycoside-modifying enzyme production suggested to us a clonal dissemination of the studied strains.