Impact of microbiology cascade reporting on antibiotic de-escalation in cefazolin-susceptible Gram-negative bacteremia.

Cascade reporting (CR) involves reporting the susceptibilities of broad-spectrum agents only when the organism is resistant to more narrow-spectrum agents. The purpose of this study is to evaluate the impact of CR on antibiotic de-escalation practices and to characterize the impact of CR on clinical outcomes. CR rules were implemented in the microbiology laboratory at Atlantic Health System (AHS) in June 2013. A retrospective chart review was conducted at two community teaching hospitals in adult patients who had a blood culture positive for a Gram-negative organism susceptible to cefazolin and who were empirically treated with broad-spectrum beta-lactam (BSBL) antibiotics. De-escalation practices were compared in the pre-CR (July 2012-December 2012) and post-CR (July 2013-December 2013) periods. The primary endpoint was the percentage of patients whose BSBL agent was de-escalated to agents listed on the post-CR antibiotic susceptibility report within 48 h of the final report. Secondary endpoints include the difference in pre-CR and post-CR periods in terms of hospital length of stay, in-hospital mortality, 30-day readmission, Clostridium difficile infections, and re-initiation of a BSBL agent within 7 days. A total of 73 patients were included; 31 in the pre-CR and 42 in the post-CR period. Patients had similar baseline characteristics. Therapy was de-escalated in 48 % of pre-CR vs 71 % of post-CR patients (p = 0.043). No significant differences were observed in secondary endpoints between patients in the pre-CR and post-CR periods. CR resulted in significant improvements in de-escalation practices without affecting safety outcomes.

Tuberculosis in elephants-a reemergent disease: diagnostic dilemmas, the natural history of infection, and new immunological tools.

Tuberculosis (TB) in elephants has been described since ancient times. However, it was not until 1996 when infection with Mycobacterium tuberculosis was identified in a herd of circus elephants that significant research into this disease began. The epidemiology and natural history of TB were unknown in elephants since there had been no comprehensive screening programs, and diagnostic techniques developed for cervidae and bovidae were of unknown value. And, while precepts of test and slaughter were the norm for cattle and deer, this was considered untenable for an endangered species. With no precedent for the treatment of TB in animals, treatment regimens for elephants were extrapolated from human protocols, which guided changes to the Guidelines for the Control of Tuberculosis in Elephants. In the absence of diagnostic testing to confirm cure in elephants, the efficacy of these treatment regimens is only beginning to be understood as treated elephants die and are examined postmortem. However, because of pressures arising from public relations related to elephant husbandry and the added considerations of TB infection in animals (whether real or imagined), sharing of information to aid in research and treatment has been problematic. Here we review the challenges and successes of the diagnosis of tuberculosis in elephants and discuss the natural history of the disease to put the work of Landolfi et al on the immunological response to tuberculosis in elephants in perspective.

Mycobacterium avium serovars 2 and 8 infections elicit unique activation of the host macrophage immune responses.

Mycobacterium avium is an opportunistic pathogen whose pathogenesis is attributed to its serovar-specific glycopeptidolipid (ssGPL), which varies among its 31 serovars. To determine if the presence and type of ssGPLs contribute to M. avium pathogenesis, we infected murine macrophages (mφs) with two M. avium wild type (wt) serovars (2 and 8) and their serovar-null strains. We examined the influence of ssGPL (presence and type) on cytokine production in non-activated (-IFN-γ) and activated (+IFN-γ) mφs, and the bacterial intra-mφ survival over a 6-day infection process. Serovar-2 infections activated TNF-α production that increased over the 6 day period and was capable of controlling the intra-mφ serovar-2 null strain. In contrast, the serovar-8 infection stimulated a strong pro-inflammatory response, but was incapable of removing the invading pathogen, maybe through IL-10 production. It was clear that the intracellular growth of serovar-null in contrast to the wt M. avium strains was easily controlled. Based on our findings and the undisputed fact that M. avium ssGPL is key to its pathogenesis, we conclude that it is not appropriate to dissect the pathogenesis of one M. avium serovar and apply those findings to other serovars.

Evidence for a clonal origin of methicillin resistance in Staphylococcus aureus.

Soon after methicillin was introduced into clinical practice in the early 1960s, resistant strains of Staphylococcus aureus (MRSA) appeared, bearing a newly acquired resistance gene, mecA, that encodes a penicillin binding protein, PBP2a. MRSA have spread throughout the world, and an investigation of the clonality of 472 isolates by DNA hybridization was performed. All 472 isolates could be divided into six temporally ordered mecA hybridization patterns, and three of these were subdivided by the chromomosomal transposon Tn554. Each Tn554 pattern occurred in association with one and only one mecA pattern, suggesting that mecA divergence preceded the acquisition of Tn554 in all cases and therefore that mecA may have been acquired just once by S. aureus.

Tuberculosis and multidrug resistance in Zambian prisons, 2000-2001.

BACKGROUND: Data on prevalence of tuberculosis (TB) and multidrug-resistant tuberculosis (MDR-TB) in Zambian prisons are lacking. METHODS: Between January 2000 and July 2001, a case-finding study was performed in 13 Zambian prisons for pulmonary TB. Prisoners were administered a questionnaire to obtain demographic information. Information regarding housing density and diet was also collected. Three consecutive first morning sputum specimens were cultured for Mycobacterium tuberculosis. Antimicrobial resistance testing was performed by the resistance ratio method. RESULTS: A total of 1080 prisoners were recruited: 1055 were males and 25 females. Sputum from 245 (22.7%) prisoners yielded M. tuberculosis, including 168 (15.6%) with smear-positive disease. Based on a total prison population of 6118, the minimal prevalence of TB was 4.0%. There was a linear relationship between the proportion of prisoners evaluated and the prevalence of TB (R(2) = 0.9366) across facilities, suggesting that the true prevalence of TB may approach 15-20%. Resistance to at least one anti-tuberculosis drug was detected for 40 (23.8%) isolates, while MDR-TB was identified for 16 (9.5%) isolates. CONCLUSION: There is a high rate of pulmonary TB in Zambian prisons, with significant rates of drug resistance and MDR-TB, highlighting the need for active surveillance and treatment programs.

Optimization of electroporation conditions for Mycobacterium avium.

Successful transformation and subsequent genetic manipulation of Mycobacterium avium requires suitable vectors, efficient transformation systems, and reliable selectable markers. A systematic analysis of the parameters involved in the transformation of M. avium was performed to optimize DNA transfer. Factors examined included the composition of the growth medium, growth medium additives, variations in washing of the bacteria prior to electroporation, and conditions of electroporation. Of the parameters assayed, the frequency of transformation (defined as the number of transformants per 10(6) transformed bacteria) showed the greatest increase with the addition of 1.5% glycine to the M. avium culture medium and the use of higher concentrations of plasmid DNA. The addition of 0.5 M sucrose to the growth medium and wash solution yielded a modest increase in transformation frequency, but more importantly afforded greater consistency of results between different batches of cells with no decrease in transformation yields following freezing and thawing. We also confirmed that gfp could be used as a selective marker for M. avium, even as a single copy integrant, and allowed for rapid discrimination between false and true transformants. Using this protocol, we were able to transform nine of 11 clinical strains of M. avium.

Clinical and bacteriologic correlates of the papG alleles among Escherichia coli strains from children with acute cystitis.

BACKGROUND: papG is the Gal(alpha1-4)Gal-specific adhesin gene of Escherichia coli P fimbriae. The three alleles of papG are associated with different receptor-binding preferences, occur in different lineages of E. coli and appear differentially associated with specific clinical syndromes, e.g. allele II with pyelonephritis and allele III with cystitis. However, no data are available regarding associations of the papG alleles with clinical outcomes. METHODS: Alleles I, II and III of papG were sought among 38 E. coli urine isolates from children with acute cystitis by a polymerase chain reaction-based assay. The papG genotype was compared with other bacterial characteristics and with response to therapy. RESULTS: papG was detected in 13 (34%) strains. It was associated positively with sfa and hly (which encode S fimbriae and hemolysin) and negatively with afa (which encodes Dr-binding adhesins). Allele II predominated over allele III (29% of strains, vs. 5%; P < 0.01). Allele II was significantly associated with serogroups O1 and O16 and with agglutination of both human and sheep erythrocytes, whereas allele III was associated with sfa, hly, serogroup 06 and preferential agglutination of sheep erythrocytes. The presence of papG predicted recurrent bacteriuria among children receiving 3-day treatment and Allele III predicted same-strain recurrence. CONCLUSIONS: These findings conflict with existing data associating allele III with cystitis, confirm and extend previous associations of papG alleles II and III with other bacterial properties and suggest that papG genotype may predict clinical outcomes.

A pseudoepidemic due to laboratory contamination deciphered by molecular analysis.

OBJECTIVE: A clinical, microbiological, and molecular analysis to identify the source of a cluster of pseudoinfections. DESIGNS: Retrospective analysis of the cases, prospective epidemiologic survey, and laboratory investigation. Molecular analysis of the isolates was performed using pulsed-field gel electrophoresis (PFGE). SETTING: A tertiary Veterans Affairs medical center. PATIENTS: Three patients admitted over a 2-week period with musculoskeletal complaints had one or more joint fluid specimens submitted for culture. In each case, anaerobic chopped meat-glucose broth (CMGB) tubes yielded one or more organisms not typically associated with septic arthritis (Enterobacter cloacae, Enterococcus faecium, Enterococcus casseliflavus, Enterococcus faecalis, Escherichia hermannii, and Pseudomonas diminuti). The first three organisms were isolated from specimens from multiple patients. Two patients had multiple positive cultures; for two patients, separate cultures yielded additional organisms on solid media. RESULTS: Laboratory investigation yielded an isolate of E faecium from 1 of 30 sham-inoculated CMGB tubes. PFGE analysis demonstrated that a single strain of E cloacae was isolated from four CMGB tubes representing all three patients, and a single strain of E faecium was isolated from CMGB tubes representing two patients and the sham-inoculated tube. CONCLUSIONS: The application of molecular typing clearly demonstrated clonality among the isolates and indicated that a common source of contamination, most likely the CMGB tubes, was responsible for these cases.

Processing postmortem specimens with C18-carboxypropylbetaine and analysis by PCR to develop an antemortem test for Mycobacterium avium infections in ducks.

Mycobacterium avium is the causative agent of the avian mycobacteriosis commonly known as avian tuberculosis (ATB). This infection causes disseminated disease, is difficult to diagnose, and is of serious concern because it causes significant mortality in birds. A new method was developed for processing specimens for an antemortem screening test for ATB. This novel method uses the zwitterionic detergent C18-carboxypropylbetaine (CB-18). Blood, bone marrow, bursa, and fecal specimens from 28 ducks and swabs of 20 lesions were processed with CB-18 for analysis by smear, culture, and polymerase chain reaction (PCR). Postmortem examination confirmed nine of these birds as either positive or highly suspect for disseminated disease. The sensitivities of smear, culture, and PCR, relative to postmortem analysis and independent of specimen type, were 44.4%, 88.9%, and 100%, respectively, and the specificities were 84.2%, 57.9%, and 15.8%, respectively. Reductions in specificity were due primarily to results among fecal specimens. However, these results were clustered among a subset of birds, suggesting that these tests actually identified birds in early stages of the disease. Restriction fragment length polymorphism mapping identified one strain of M. avium (serotype 1) that was isolated from lesions, bursa, bone marrow, blood, and feces of all but three of the culture-positive birds. In birds with confirmed disease, blood had the lowest sensitivity and the highest specificity by all diagnostic methods. Swabs of lesions provided the highest sensitivity by smear and culture (33.3% and 77.8%, respectively), whereas fecal specimens had the highest sensitivity by PCR (77.8%). The results of this study indicate that processing fecal specimens with CB-18, followed by PCR analysis, may provide a valuable first step for monitoring the presence of ATB in birds.

Population pharmacokinetics of pyrazinamide in elephants.

This study was undertaken to characterize the population pharmacokinetics (PK), therapeutic dose, and preferred route of administration for pyrazinamide (PZA) in elephants. Twenty-three African (Loxodonta africana) and Asian (Elephas maximus) elephants infected with or in contact with others culture positive for Mycobacterium tuberculosis were dosed under treatment conditions. PZA was dosed daily at 20-30 mg/kg via oral (fasting or nonfasting state) or rectal (enema or suppository) administration. Blood samples were collected 0-24 h postdose. Population PK was estimated using nonlinear mixed effect modeling. Drug absorption was rapid with T(max) at or before 2 h regardless of the method of drug administration. C(max) at a mean dose of 25.6 (+/-4.6) mg/kg was 19.6 (+/-9.5 microg/mL) for PZA given orally under fasting conditions. Under nonfasting conditions at a mean dose of 26.1 +/- 4.2 mg/kg, C(max) was 25% (4.87 +/- 4.89 microg/mL) and area under concentration curve (AUC) was 30% of the values observed under fasting conditions. Mean rectal dose of 32.6 +/- 15.2 mg/kg yielded C(max) of 12.3 +/- 6.3 microg/mL, but comparable AUC to PZA administered orally while fasting. Both oral and rectal administration of PZA appeared to be acceptable and oral dosing is preferred because of the higher C(max) and lower inter-subject variability. A starting dose of 30 mg/kg is recommended with drug monitoring between 1 and 2 h postdose. Higher doses may be required if the achieved C(max) values are below the recommended 20-50 microg/mL range.

Population pharmacokinetics of isoniazid in the treatment of Mycobacterium tuberculosis among Asian and African elephants (Elephas maximus and Loxodonta africana).

We recently described the clinical presentation and treatment of 18 elephants from six herds infected with TB. Treatment protocols and methods varied between herds to include both oral and rectal dosing using multiple drug doses and formulations. In this paper we present information regarding the pharmacokinetics (PK) of isoniazid (INH) in elephants and provide suggestions regarding initial treatment regimens. Forty-one elephants received INH daily by either oral or rectal administration with different formulations. Population PK analysis was performed using Non-linear Mixed Effect Modeling (NONMEM). Results of oral administration indicated that compared with premixed INH solution, the drug exposure was highest with a suspension prepared freshly with INH powder. When INH was concomitantly given as an admixture over food, Tmax was delayed and variability in drug absorption was significantly increased. Compared with oral administration, similar drug exposures were found when INH was dosed rectally. The data generated suggest that a starting dose of 7.5 mg/kg of INH is appropriate for initial TB treatment in elephants when premixed solution is administered directly into the oropharynx or rectal vault and 4 mg/kg are when INH is administered following immediate suspension from powdered form.

Clinical syndromes associated with adult pneumococcal cellulitis.

Streptococcus pneumoniae is an uncommonly recognized etiology of cellulitis in adults. A review of the literature uncovered 30 cases of pneumococcal skin infection in adults. Typically, all patients with pneumococcal cellulitis had an underlying chronic illness, or were immunocompromised by drug or alcohol abuse. Pneumococcal cellulitis presents as two distinctive clinical syndromes: one with extremity involvement in individuals with diabetes and substance abuse; and a second involving the head, neck and upper torso in individuals with systemic lupus erythematosis, nephrotic syndrome and hematologic disorders. For each there are statistically significant associations between the location of pneumococcal cellulitis and underlying clinical disorders. In contrast to other common bacterial etiologies, pneumococcal cellulitis is frequently associated with blood stream invasion, tissue necrosis and suppurative complications. Patients often require surgical interventions and prolonged hospitalizations. A high degree of suspicion and early aggressive management is needed for those presenting with cellulitis characterized by bullae and violaceous color.

Polymicrobial and recurrent bacteremia with Shigella in a patient with AIDS.

Shigella gastroenteritis is uncommon among HIV seropositive patients and may be complicated in some patients by bacteremia; S. flexneri being the most frequently detected serogroup. While recurrent Salmonella bacteremia is common among HIV-seropositive patients, recurrent Shigella bacteremia is not. We report here an HIV-seropositive patient with Shigella gastroenteritis, polymicrobial bacteremia due to S. flexneri and S. boydii, and recurrent gastroenteritis and bacteremia with S. boydii. Relapsing infection with the same strain of S. boydii was determined using pulsed field gel electrophoresis. Thus, HIV-seropositive patients who develop Shigella infections may require prolonged treatment and/or suppressive therapy, similar to those infected with Salmonella. Patients who develop recurrent disease should be suspected as having polymicrobial bacteremia since the incidence of this may be underestimated among patients with AIDS, particularly those with concurrent gastroenteritis.

Recurrent Escherichia coli bacteremia.

Escherichia coli is the most common gram-negative organism associated with bacteremia. While recurrent E. coli urinary tract infections are well-described, recurrent E. coli bacteremia appears to be uncommon, with no episodes noted in multiple series of patients with gram-negative bacteremias. We report on 5 patients with recurrent bloodstream infections identified from a series of 163 patients with E. coli bacteremia. For each patient, the isolates from each episode were analyzed by pulsed-field gel electrophoresis (PFGE) and ribotyping and for the presence of E. coli virulence factors. For each of four patients, the index and recurrent episodes of bacteremia represented the same strain as defined by PFGE, and the strains were found to carry one or more virulence factors. The remaining patient, with two episodes of bloodstream infection separated by a 4-year interval, was infected with two isolates that did not carry any virulence factors and that were clonally related by ribotype analysis but differed by PFGE. All five patients had either a local host defense defect (three patients) or impaired systemic defenses (one patient) or both (one patient). Thus, recurrent E. coli bacteremia is likely to represent a multifactorial process that occurs in patients with impaired host defenses who are infected with virulent isolates.

Clonal distribution of the three alleles of the Gal(alpha1-4)Gal-specific adhesin gene papG among Escherichia coli strains from patients with bacteremia.

The three alleles of papG, the Gal(alpha1-4)Gal-specific adhesin gene of Escherichia coli P fimbriae, were detected in 101 (54%) of 187 E. coli bacteremia isolates from Boston, Long Beach, California, and Nairobi, Kenya, by a polymerase chain reaction-based assay, and their distribution was compared with clonal structure and other bacterial and host characteristics. Allele II predominated overall (57% of papG+ strains, vs. 32% for III and 2% for I). Allele distribution differed significantly between centers. Alleles segregated according to enzyme electrophoretic lineage and O serogroup in patterns suggesting both vertical and horizontal transmission and emergence within E. coli in the temporal sequence II-->III-->I. Strains containing allele III preferentially agglutinated sheep erythrocytes, whereas strains containing allele II preferentially agglutinated human erythrocytes. Allele III was associated with hly, sfa, multiple copies of the pap operon, and cluster III of the phylogenetic tree. These findings indicate that among bacteremia isolates of E. coli, the three papG alleles exhibit distinctive associations with evolutionary lineages, hemagglutination phenotypes, and non-pap virulence properties.

Persistent and relapsing infections associated with small-colony variants of Staphylococcus aureus.

Small-colony variants (SCVs) of Staphylococcus aureus were cultured from five patients with persistent and relapsing infections. All five SCV strains were nonhemolytic and nonpigmented and grew very slowly on routine culture media in an ambient atmosphere. In several instances, these phenotypic characteristics led to the initial misidentification of the organisms in the clinical microbiology laboratory. All four strains available for further analysis were shown to be auxotrophs that reverted to normal growth and morphology in the presence of menadione, hemin, and/or a CO2 supplement. Similarly, these isolates were resistant to aminoglycosides under routine conditions but susceptible in the presence of the metabolic supplements. For two patients, the large and small colony forms isolated concurrently were indistinguishable when analyzed by pulsed field gel electrophoresis and thus represented phenotypic variants within individual clones. We propose a model relating the phenotypic characteristics of S. aureus SCVs with the clinical pattern of persistent and relapsing infection.

Related strains of Mycobacterium avium cause disease in children with AIDS and in children with lymphadenitis.

Sequence analysis of the ribosomal internal transcribed spacer of 56 Mycobacterium avium complex isolates from pediatric patients with AIDS or lymphadenitis revealed (similar to the situation in adults) that the closely related Mav-B and Mav-A sequevars caused the vast majority of disease. IS1245 restriction fragment-polymorphism analysis and pulsed-field gel electrophoresis revealed sets of isolates with closely related patterns among strains from patients in the Boston area and among isolates from Los Angeles and Miami patients. The finding of related strains that cause disease in epidemiologically unrelated patients is most consistent with one of two hypotheses: (1) a limited subset of M. avium strains is more virulent and therefore more likely to cause disease in humans, or (2) pathogenic strains are more prevalent in the environment.

Hemolysin as a virulence factor for systemic infection with isolates of Mycobacterium avium complex.

Isolates of the Mycobacterium avium complex were examined for hemolysin expression. Only invasive isolates of M. avium were observed to be hemolytic (P < 0.001), with activity the greatest for isolates of serovars 4 and 8. Thus, M. avium hemolysin appears to represent a virulence factor necessary for invasive disease.

Mycobacterium tuberculosis infection as a zoonotic disease: transmission between humans and elephants.

Between 1994 and 1996, three elephants from an exotic animal farm in Illinois died of pulmonary disease due to Mycobacterium tuberculosis. In October 1996, a fourth living elephant was culture-positive for M. tuberculosis. Twenty-two handlers at the farm were screened for tuberculosis (TB); eleven had positive reactions to intradermal injection with purified protein derivative. One had smear-negative, culture-positive active TB. DNA fingerprint comparison by IS6110 and TBN12 typing showed that the isolates from the four elephants and the handler with active TB were the same strain. This investigation indicates transmission of M. tuberculosis between humans and elephants.

Ongoing horizontal and vertical transmission of virulence genes and papA alleles among Escherichia coli blood isolates from patients with diverse-source bacteremia.

The phylogenetic distributions of multiple putative virulence factors (VFs) and papA (P fimbrial structural subunit) alleles among 182 Escherichia coli blood isolates from patients with diverse-source bacteremia were defined. Phylogenetic correspondence among these strains, the E. coli Reference (ECOR) collection, and other collections of extraintestinal pathogenic E. coli (ExPEC) was assessed. Although among the 182 bacteremia isolates phylogenetic group B2 predominated, exhibited the greatest concentration of individual VFs, and contained the largest number of familiar virulent clones, other phylogenetic groups exhibited greater concentrations of certain VFs than did group B2 and included several additional virulent clones. Certain of the newly detected VF genes, e.g., fyuA (yersiniabactin; 76%) and focG (F1C fimbriae; 25%), were as prevalent or more prevalent than their more familiar traditional counterparts, e.g., iut (aerobactin; 57%) and sfaS (S fimbriae; 14%), thus possibly offering additional useful targets for preventive interventions. Considerable diversity of VF profiles was observed at every level within the phylogenetic tree, including even within individual lineages. This suggested that many different pathways can lead to extraintestinal virulence in E. coli and that the evolution of ExPEC, which involves extensive horizontal transmission of VFs and continuous remodeling of pathogenicity-associated islands, is a highly active, ongoing process.