Effect of on-farm poultry litter composting processes on physicochemical, biological, and toxicological parameters and reduction of antibiotics and antibiotic-resistant Escherichia coli.

Poultry litter is a valuable source of nutrients for crop production, but its use in agriculture can lead to environmental and public health concerns due to the presence of pollutants, antibiotic-resistant bacteria (ARB) and antibiotic-resistant genes (ARGs). We compared the effect of different on-farm poultry litter composting processes on physicochemical, biological, and toxicological parameters, as well as on the occurrence of antibiotics and resistant Escherichia coli. The composting treatments consisted of passively-aerated piles C:N = 19 (PAC19), mechanically-aerated piles C:N = 19 (MAC19), and mechanically-aerated piles C:N = 30 (MAC30). Poultry litter composting led to a significant reduction of antibiotic residues, enteroparasites and antibiotic resistant E. coli. The conditions of the process, such as extra C source and mechanical aeration influence the quality of the final product. MAC19 is a low-cost effective method to reduce the potential risks associated with poultry litter use in agriculture and produce good quality compost.

Detection of toxigenic Clostridioides --#1;Clostridium--#3; difficile: Usefulness of two commercially available enzyme immunoassays and a PCR assay on stool samples and stool isolates / Detección de Clostridioides --#1;Clostridium--#3; difficile toxigénico: utilidad de dos métodos de enzimoinmunoensayo comerciales y una PCR en las muestras de materia fecal y en los respectivos aislamientos

The best laboratory diagnostic approach to detect Clostridioides --#1;Clostridium--#3; difficile infection (CDI) is a subject of ongoing debate. With the aim of evaluating four laboratory diagnostic methods, 250 unformed stools from patients with suspected CDI submitted to nine medical center laboratories from November 2010 to December 2011, were studied using: (1) an immunochromatographic rapid assay test that combines the qualitative determination of glutamate dehydrogenase (GDH) plus toxins A and B (QAB), the CDIFF QUIK CHEK COMPLETE assay; (2) an enzyme immunoassay for qualitative determination of toxins A and B, the RIDASCREENTC. difficile Toxin A/B assay (RAB); (3) a PCR for the toxin B gene assay (PCR); and (4) the toxigenic culture (TC).C. difficile isolates from direct toxin negative stools by QAB, RAB and PCR were evaluated for toxigenicity by the same direct tests, in order to assess the contribution of the TC (QAB-TC, RAB-TC, PCR-TC). A combination of the cell culture cytotoxicity neutralization assay (CCCNA) in stools, and the same assay on isolates from direct negative samples (CCCNA-TC) was considered the reference method (CCCNA/CCCNA-TC). Of the 250 stools tested, 107 (42.8%) were positive by CCCNA/CCCNA-TC. The GDH and PCR/PCR-TC assays were the most sensitive, 91.59% and 87.62%, respectively. The QAB, RAB, QAB/QAB-TC and RAB/RAB-TC had the highest specificities, ca. 95%. A negative GDH result would rule out CDI, however, its low positive likelihood ratio (PLR) of 3.97 indicates that a positive result should always be complemented with the detection of toxins. If the RAB, QAB, and PCR assays do not detect toxins from direct feces, the toxigenic culture should be performed. In view of our results, the most accurate and reliable methods to be applied in a clinical microbiology laboratory were the QAB/QAB-TC, and RAB/RAB-TC, with PLRs >10 and negative likelihood ratios <0.30.

El mejor procedimiento para realizar el diagnóstico de laboratorio de la infección causada por Clostridioides --#1;Clostridium--#3; difficile (ICD) es aún objeto de debate. Con el fin de evaluar cuatro métodos diagnósticos de laboratorio, se estudiaron 250 muestras de heces diarreicas provenientes de pacientes con sospecha de ICD remitidas a los laboratorios de nueve centros médicos entre noviembre de 2010 y diciembre de 2011. Dichas muestras se analizaron mediante los siguientes métodos:1) un ensayo rápido inmunocromatográfico que combina la detección cualitativa de la glutamato deshidrogenasa (GDH) y de las toxinas Ay B (QAB), CDIFF QUIK CHEK COMPLETE;2) un enzimoinmunoanálisis para la determinación cualitativa de las toxinas A/B, RIDASCREENTC. difficile Toxin A/B (RAB);3) un método molecular basado en PCR para la detección del gen que codifica la toxina B (PCR) y 4) el cultivo toxigénico (TC). Como método de referencia se utilizó la combinación del ensayo de citotoxicidad sobre cultivo de células con la neutralización de toxina mediante anticuerpo específico en los filtrados de las heces (CCCNA) y el mismo método en sobrenadantes de aislamientos de C. difficile (CCCNA-TC). La toxigenicidad de las cepas aisladas de muestras directas negativas con QAB, RAB y PCR se evaluó con los mismos métodos, con el propósito de detectar la contribución del TC (QAB-TC, RAB-TC, PCR-TC). De las 250 muestras estudiadas, 107 (42,8%) fueron positivas por CCCNA/CCCNA-TC. Los métodos GDH y PCR/PCR-TC fueron los más sensibles: 91,59 y 87,62%, respectivamente. Los métodos QAB, RAB, QAB/QAB-TC y RAB/RAB-TC mostraron las mayores especificidades, del 95%, aproximadamente. Un resultado negativo para GDH excluiría la ICD, pero su baja razón de verosimilitud positiva (PLR), que fue 3,97, indica que un resultado positivo debe complementarse con la detección de toxinas. Cuando no se detectan toxinas directas por RAB, QAB ni PCR, debería realizarse el TC. De acuerdo con nuestros resultados, los métodos más precisos y confiables para ser aplicados en un laboratorio de microbiología clínica son QAB/QAB-TC y RAB/RAB-TC, con una PLR> 10 y una razón de verosimilitud negativa < 0,30.

Detection of toxigenic Clostridioides [Clostridium] difficile: Usefulness of two commercially available enzyme immunoassays and a PCR assay on stool samples and stool isolates.

The best laboratory diagnostic approach to detect Clostridioides [Clostridium] difficile infection (CDI) is a subject of ongoing debate. With the aim of evaluating four laboratory diagnostic methods, 250 unformed stools from patients with suspected CDI submitted to nine medical center laboratories from November 2010 to December 2011, were studied using: (1) an immunochromatographic rapid assay test that combines the qualitative determination of glutamate dehydrogenase (GDH) plus toxins A and B (QAB), the CDIFF QUIK CHEK COMPLETE assay; (2) an enzyme immunoassay for qualitative determination of toxins A and B, the RIDASCREEN™ C. difficile Toxin A/B assay (RAB); (3) a PCR for the toxin B gene assay (PCR); and (4) the toxigenic culture (TC). C. difficile isolates from direct toxin negative stools by QAB, RAB and PCR were evaluated for toxigenicity by the same direct tests, in order to assess the contribution of the TC (QAB-TC, RAB-TC, PCR-TC). A combination of the cell culture cytotoxicity neutralization assay (CCCNA) in stools, and the same assay on isolates from direct negative samples (CCCNA-TC) was considered the reference method (CCCNA/CCCNA-TC). Of the 250 stools tested, 107 (42.8%) were positive by CCCNA/CCCNA-TC. The GDH and PCR/PCR-TC assays were the most sensitive, 91.59% and 87.62%, respectively. The QAB, RAB, QAB/QAB-TC and RAB/RAB-TC had the highest specificities, ca. 95%. A negative GDH result would rule out CDI, however, its low positive likelihood ratio (PLR) of 3.97 indicates that a positive result should always be complemented with the detection of toxins. If the RAB, QAB, and PCR assays do not detect toxins from direct feces, the toxigenic culture should be performed. In view of our results, the most accurate and reliable methods to be applied in a clinical microbiology laboratory were the QAB/QAB-TC, and RAB/RAB-TC, with PLRs >10 and negative likelihood ratios <0.30.

Transposons and integrons in colistin-resistant clones of Klebsiella pneumoniae and Acinetobacter baumannii with epidemic or sporadic behaviour.

Multiple transposons, integrons and carbapenemases were found in Klebsiella pneumoniae colistin-resistant isolates as well as a genomic resistance island of the AbaR type in Acinetobacter baumannii colistin-resistant isolates from different hospitals from Buenos Aires City. PFGE analysis showed a polyclonal dissemination of antimicrobial resistance mechanisms among K. pneumoniae isolates, while in A. baumannii isolates the epidemic clone 1 from South America was found. Resistance determinants associated with horizontal gene transfer are contributing to the evolution to pandrug resistance in both epidemic and sporadic clones.

First national survey of antibiotic susceptibility of the Bacteroides fragilis group: emerging resistance to carbapenems in Argentina.

The antibiotic susceptibility rates of 363 clinical Bacteroides fragilis group isolates collected from 17 centers in Argentina during the period from 2006 to 2009 were as follows: piperacillin-tazobactam, 99%; ampicillin-sulbactam, 92%; cefoxitin, 72%; tigecycline, 100%; moxifloxacin, 91%; and clindamycin, 52%. No metronidazole resistance was detected in these isolates during this time period. Resistance to imipenem, doripenem, and ertapenem was observed in 1.1%, 1.6%, and 2.3% of B. fragilis group strains, respectively. B. fragilis species showed a resistance profile of 1.5% to imipenem, 1.9% to doripenem, and 2.4% to ertapenem. This is the first report of carbapenem resistance in Argentina. The cfiA gene was present in 8 out of 23 isolates, all of them belonging to the B. fragilis species and displaying reduced susceptibility or resistance to carbapenems (MICs ≥ 4 µg/ml). Three out of eight cfiA-positive isolates were fully resistant to carbapenems, while 5 out of 8 isolates showed low-level resistance (MICs, 4 to 8 µg/ml). The inhibition by EDTA was a good predictor of the presence of metallo-ß-lactamases in the fully resistant B. fragilis strains, but discrepant results were observed for low-level resistant isolates. B. fragilis was more susceptible to antimicrobial agents than other Bacteroides species. Bacteroides vulgatus species was the most resistant to ampicillin-sulbactam and piperacillin-tazobactam, and B. thetaiotaomicron/ovatus strains showed the highest level of resistance to carbapenems, with an unknown resistance mechanism. B. vulgatus and the uncommon non-Bacteroides fragilis species were the most resistant to moxifloxacin, showing an overall resistance rate of 15.1%.

[First Argentine consensus guidelines for in vitro antimicrobial susceptibility testing of clinically relevant anaerobic bacteria in humans/ Anaerobic Subcommittee of the Asociación Argentina de Microbiología]. / Primer consenso argentino para el estudio de la sensibilidad in vitro a los antimicrobianos de las bacterias anaerobias de importancia clínica en humanos.

Through time, anaerobic bacteria have shown good susceptibility to clinically useful antianaerobic agents. Nevertheless, the antimicrobial resistance profile of most of the anaerobic species related to severe infections in humans has been modified in the last years and different kinds of resistance to the most active agents have emerged, making their effectiveness less predictable. With the aim of finding an answer and for the purpose of facilitating the detection of anaerobic antimicrobial resistance, the Anaerobic Subcommittee of the Asociación Argentina de Microbiología developed the First Argentine consensus guidelines for in vitro antimicrobial susceptibility testing of clinically relevant anaerobic bacteria in humans. This document resulted from the compatibilization of the Clinical and Laboratory Standards Institute recommendations, the international literature and the work and experience of the Subcommittee. The Consensus document provides a brief taxonomy review, and exposes why and when anaerobic antimicrobial susceptibility tests should be conducted, and which antimicrobial agents can be used according to the species involved. The recommendations on how to perform, read and interpret in vitro anaerobic antimicrobial susceptibility tests with each method are exposed. Finally, the antibiotic susceptibility profile, the classification of antibiotics according to their in vitro activities, the natural and acquired mechanisms of resistance, the emerging resistance and the regional antibiotic resistance profile of clinically relevant anaerobic species are shown.

Polyclonal spread of bla(OXA-23) and bla(OXA-58) in Acinetobacter baumannii isolates from Argentina.

BACKGROUND: In order to study the enzymatic carbapenem resistance mechanisms in Acinetobacter baumannii isolates from Argentina, we performed molecular characterization on 41 epidemiologically unrelated strains isolated from 1995 to 2006 with diminished susceptibilities to imipenem and meropenem. METHODOLOGY: Acinetobacter baumannii isolates were identified with the ARDRA technique. The total genomic DNA was used to detect each carbapenem beta-lactamase gene described so far in this species and those insertion sequences usually associated to carbapenem beta-lactamase genes (ISAba1, 2, 3, 4 and IS18) by the PCR technique with specific primers. RESULTS: 26 out of 41 Acinetobacter baumannii isolates with diminished susceptibilities to carbapenems harboured the bla(OXA-23) gene. The bla(OXA-58) was detected in 13 out of 41 isolates. ISAba1 was always located upstream bla(OXA-23). All isolates containing the bla(OXA-58) gene showed ISAba3 downstream of the carbapenemase, while 4 isolates had a second copy of the ISAba3 upstream of the gene. CONCLUSION: Enzymatic carbapenem resistance in Acinetobacter baumannii was found in 88% of 41 non-epidemiologically-related strains mediated by the polyclonal spread of the bla(OXA-23) and bla(OXA-58) genes. The genetic structures surrounding the oxacillinase genes found in our bacterial isolates revealed a particular epidemiology in our geographical region. This data suggests the need of local molecular surveillance to help control multirresistance Acinetobacter baumannii infections.

Pneumococcal bacteremia in a single center in Argentina.

OBJECTIVE: To determine the clinical and microbiologic characteristics of pneumococcal bacteremia at Sanatorio Mitre, Buenos Aires, Argentina. METHODS: One-hundred-and-seven episodes of pneumococcal bacteremia were prospectively analyzed from 1993 to 1998. Demographics, clinical and microbiological variables were studied. RESULTS: Eighty-one patients (76%) were adults and 26 children (24%). Most cases (98%) were acquired in the community. Seventy-nine patients (74%) had at least one underlying condition. The primary source of bacteremia was found in 91 patients (85%), the lungs being the most common source. Streptococcus pneumoniae was isolated from one sterile site other than the primary focus in 25 patients (23%). Eighty-five (79%) of the Streptococcus pneumoniae were susceptible to penicillin and 22 (21%) showed intermediate or high resistance to penicillin and 2% were additionally resistant to ceftriaxone. Initial antimicrobial therapy was appropriate in 95% of the cases. The overall mortality was 21%, however adults admitted to the intensive care unit (ICU) had higher mortality (81%). No patients under 14 years old died. Multivariate analysis showed that age and recovery of the organisms from a sterile site other than the primary focus were statistically significant predictors of mortality. CONCLUSION: Bacteremic pneumococcal infections continue to be an important worldwide problem causing morbidity and high mortality despite supportive care and appropriate antimicrobial therapy.

Identification of an epidemic carbapenem-resistant Acinetobacter baumannii strain at hospitals in Buenos Aires City.

To identify epidemic Acinetobacter baumannii (AB) clones, 38 carbapenem-resistant AB isolates from 5 hospitals were analyzed. Macrorestriction classified 24 isolates as clone IV, susceptibility pattern clustering analysis grouped almost all of them together, and they were uniformly biotype 8. Clone IV was present at all 5 hospitals, so that it represents a carbapenem-resistant AB strain with epidemic behavior.