Planning to halve Gram-negative bloodstream infection: getting to grips with healthcare-associated Escherichia coli bloodstream infection sources.

BACKGROUND: A thorough understanding of the local sources, risks, and antibiotic resistance for Escherichia coli bloodstream infection (BSI) is required to focus prevention initiatives and therapy. AIM: To review the sources and antibiotic resistance of healthcare-associated E. coli BSI. METHODS: Sources and antibiotic resistance profiles of all 250 healthcare-associated (post 48 h) E. coli BSIs that occurred within our secondary and tertiary care hospital group from April 2014 to March 2017 were reviewed. Epidemiological associations with urinary source, gastrointestinal source, and febrile neutropenia-related BSIs were analysed using univariable and multivariable binary logistic regression models. FINDINGS: E. coli BSIs increased 9% from 4.0 to 4.4 per 10,000 admissions comparing the 2014/15 and 2016/17 financial years. Eighty-nine cases (36%) had a urinary source; 30 (34%) of these were classified as urinary catheter-associated urinary tract infections (UTIs). Forty-five (18%) were related to febrile neutropenia, and 38 (15%) had a gastrointestinal source. Cases were rarely associated with surgical procedures (11, 4%) or indwelling vascular devices (seven, 3%). Female gender (odds ratio: 2.3; 95% confidence interval: 1.2-4.6) and older age (1.02; 1.00-1.05) were significantly associated with a urinary source. No significant associations were identified for gastrointestinal source or febrile neutropenia-related BSIs. Forty-seven percent of the isolates were resistant to ciprofloxacin, 37% to third-generation cephalosporins, and 22% to gentamicin. CONCLUSION: The gastrointestinal tract and febrile neutropenia together accounted for one-third of E. coli BSI locally but were rare associations nationally. These sources need to be targeted locally to reduce an increasing trend of E. coli BSIs.

Sequence assembly.

Despite the rapidly increasing number of sequenced and re-sequenced genomes, many issues regarding the computational assembly of large-scale sequencing data have remain unresolved. Computational assembly is crucial in large genome projects as well for the evolving high-throughput technologies and plays an important role in processing the information generated by these methods. Here, we provide a comprehensive overview of the current publicly available sequence assembly programs. We describe the basic principles of computational assembly along with the main concerns, such as repetitive sequences in genomic DNA, highly expressed genes and alternative transcripts in EST sequences. We summarize existing comparisons of different assemblers and provide a detailed descriptions and directions for download of assembly programs at: http://genome.ku.dk/resources/assembly/methods.html.

SNP-finding in pig mitochondrial ESTs.

The Sino-Danish pig genome project produced 685 851 ESTs (Gorodkin et al. 2007), of which 41 499 originated from the mitochondrial genome. In this study, the mitochondrial ESTs were assembled, and 374 putative SNPs were found. Chromatograms for the ESTs containing SNPs were manually inspected, and 112 total (52 non-synonymous) SNPs were found to be of high confidence (five of them are close to disease-causing SNPs in humans). Nine of the high-confidence SNPs were tested experimentally, and eight were confirmed. The SNPs can be accessed online at http://pigest.ku.dk/more/mito.

Detection of Rickettsia, Borrelia, and Bartonella in Carios kelleyi (Acari: Argasidae).

Carios kelleyi (Colley & Kohls 1941), a tick associated with bats and bat habitats, has been reported to feed on humans, but there is little published data regarding the presence of vector-borne pathogens in these ticks. C. kelleyi nymphs and adults were collected from residential and community buildings in Jackson County, Iowa, and tested by polymerase chain reaction for Rickettsia, Borrelia, Bartonella, Coxiella, and Anaplasma. Rickettsia DNA was detected in 28 of 31 live ticks. Sequences of the 17-kDa and rOmpA genes suggest that this agent is a novel spotted fever group Rickettsia. Transstadial and transovarial transmission of this Rickettsia were demonstrated. The flagellin gene of a Borrelia, closely related to B. turicatae, was detected in one of 31 live ticks. The 16S-23S intergenic spacer region of Bartonella henselae also was detected in one of 31 live ticks. Coxiella or A. phagocytophilum DNA were not detected in these ticks.

Detection of human blood in the bat tick Carios (Ornithodoros) kelleyi (Acari: Argasidae) in Iowa.

The argasid tick Carios (Ornithodoros) kelleyi Cooley & Kohls is a common ectoparasite of bats and has been found in massive numbers in homes with associated bat colonies in eastern Iowa. This tick feeds nearly exclusively on bats in nature. Several inhabitants of infested homes complained of "bug bites" at night while asleep that may have resulted in erythematous, edematous, urticaric skin lesions and constitutional signs and symptoms. We provide laboratory evidence that a single, engorged C. kelleyi nymph contained host blood from a human female. The clinical implications of our findings are intriguing but unclear.

Trans-sodium crocetinate restores blood pressure, heart rate, and plasma lactate after hemorrhagic shock.

BACKGROUND: Trans-sodium crocetinate (TSC) has been shown to increase oxygen consumption during hemorrhagic shock. The current study was done to determine the effect of TSC on other parameters such as blood pressure, heart rate, blood pH, and lactate. METHODS: A rat model of hemorrhagic shock was used, in which a constant volume of blood is removed. RESULTS: TSC increased mean arterial blood pressure from a value (immediately after hemorrhage) of 35 mm Hg to a value of 75 mm Hg, and all treated animals survived. In contrast, blood pressure in control animals decreased, with most dying soon after the hemorrhage. TSC also lessened the tachycardia which resulted from the hemorrhage. Blood pH did not decrease as much when TSC was given, and plasma lactate levels were greatly reduced. CONCLUSION: It would appear that TSC is a promising initial treatment for hemorrhagic shock.

A national laboratory network for bioterrorism: evolution from a prototype network of laboratories performing routine surveillance.

The need for an enhanced network of laboratories to respond to a bioterrorism attack has been realized. Therefore, the Association of Public Health Laboratories and the Centers for Disease Control are developing a system involving civilian public health and private laboratories that builds on the existing network for routine disease surveillance. It is anticipated that most bioterrorist attacks will not be immediately recognized, so increased laboratory capabilities and communications are necessary. The laboratory network has four categories with different biosafety levels assigned to clearly delineate the correct referral route. Improving communications through World Wide Web-based systems will allow test results, surge capacity, and training and identification algorithms to be shared instantly. There are plans to expand the network to include standard public health surveillance and emerging infectious diseases.

Ehrlichia chaffeensis antibodies in white-tailed deer, Iowa, 1994 and 1996.

Surveillance of 2,277 white-tailed deer for antibodies against Ehrlichia chaffeensis in Iowa showed seropositivity rates of 12.5% in 1994 and 13.9% in 1996. From 1994 to 1996, the estimated number of seropositive deer increased to 54,701 (28%). The increasing deer population and expanding tick distribution may increase risk for human monocytic ehrlichiosis.

Hepatitis E virus infection prevalence among selected populations in Iowa.

BACKGROUND: HEV causes an enteric infectious disease endemic in developing areas with hot climate. A case of endogenous HEV infection has been reported in the US. Recently, HEV-like virus was isolated from swine in Iowa. Swine production is a major industry in Iowa with the potential for human exposure to swine in and around industrial and family farm operations. OBJECTIVE: The study objective was to determine whether individuals in Iowa are exposed to HEV. STUDY DESIGN: Anti-HEV antibody prevalence in four selected Iowa populations was determined. Sera were collected from 204 patients with non-A, non-B, non-C hepatitis (non-A-C); 87 staff members of the Department of Natural Resources (DRN); 332 volunteer blood donors in 1989; and 111 volunteer blood donors in 1998. All sera were tested for anti-human HEV IgM and IgG by ELISA with confirmation of positivity by a peptide neutralization test. RESULTS: Both the patients with non-A, non-B, non-C hepatitis (4.9%) and the healthy field workers from the Iowa DNR (5.7%) showed significantly higher prevalence of anti-HEV IgG antibodies compared to normal blood donor sera collected in 1998 (P < 0.05). CONCLUSIONS: Human HEV or a HEV-like agent circulates in the Iowa geographical area. At-risk human populations with occupational exposure to wild animals and environmental sources of domestic animal wastes or with unexplained hepatitis have increased seroprevalence of HEV antibodies.

Comparison of PCR, culture, and direct fluorescent-antibody testing for detection of Bordetella pertussis.

We prospectively compared the performance of culture, direct fluorescent-antibody testing (DFA), and an in-house-developed PCR test targeting the repeated insertion sequence IS481 for the detection of Bordetella pertussis in nasopharyngeal swab specimens. We tested 319 consecutive paired specimens on which all three tests were performed. A total of 59 specimens were positive by one or more tests. Of these, 5 were positive by all three tests, 2 were positive by culture and PCR, 16 were positive by PCR and DFA, 28 were positive by PCR only, and 8 were positive by DFA only. Any specimen positive by culture was considered to be a true positive, as were specimens positive by both PCR and DFA. Specimens positive only by PCR or DFA were considered discrepant, and their status was resolved by review of patient histories. Patients with symptoms meeting the Centers for Disease Control and Prevention clinical case definition for pertussis and who had a specimen positive by PCR or DFA were considered to have true B. pertussis infections. Of the 28 patients positive by PCR only, 20 met the clinical case definition for pertussis, while 3 of the 8 patients positive by DFA only met the clinical case definition. After resolution of the status of discrepant specimens, the sensitivity, specificity, positive predictive value, and negative predictive value were 15.2, 100, 100, and 87.5%, respectively, for culture; 93.5, 97.1, 84.3, and 98.9%, respectively, for PCR; and 52.2, 98.2, 82.8, and 92.4%, respectively, for DFA. The actual positive predictive value of PCR was probably greater, as several PCR-positive patients who did not meet the clinical case definition had symptoms consistent with typical or atypical pertussis. PCR is a sensitive and specific method for the detection of B. pertussis.

Insertion element IS3-based PCR method for subtyping Escherichia coli O157:H7.

An Escherichia coli O157:H7 subtyping method based on PCR amplification of variable DNA sequences between the repetitive element IS3 was developed. Template DNA was prepared by boiling cells in Chelex. Two separate IS3 PCR amplifications were performed for each isolate: one with a single primer (primer IS3A) and one with two primers (primers IS3A and IS3B). The IS3 PCR subtyping method was applied to 35 epidemiologically related and unrelated E. coli O157:H7 isolates that had been previously characterized by pulsed-field gel electrophoresis (PFGE). PFGE identified 25 different subtypes (difference of one or more bands). PCR with single primer IS3A and primer pair IS3A-IS3B identified 6 and 14 different subtypes, respectively. By combining the results of the two PCR amplifications, 15 different IS3 PCR subtypes were identified. While not as sensitive as PFGE, IS3 PCR subtyping grouped all outbreak-related isolates. IS3 PCR banding patterns were reproducible between amplifications and between subcultures. IS3 PCR could serve as a simple, rapid screening method for the identification of unrelated E. coli O157:H7 isolates.

Polymerase chain reaction typing of nonviable Mycobacterium tuberculosis isolates.

Repetitive element polymerase chain reaction (PCR) typing was applied to two Mycobacterium tuberculosis isolates for which both viable and nonviable cultures were available. DNA extracted from the nonviable cultures and from fresh subcultures of the viable cultures was amplified with primers directed against the insertion sequence IS6110 and the polymorphic GC-rich repetitive sequence. For both isolates, the nonviable cultures generated PCR-banding patterns identical to those generated by the fresh subcultures.

Herpes simplex viremia: report of eight pediatric cases and review of the literature.

Bloodstream infection due to herpes simplex virus (HSV) is rare in the immunocompetent host but may be important in the pathogenesis of disseminated HSV infection in the immunocompromised patient. Using a simple blood-culture method, we detected herpes simplex viremia in eight immunologically compromised or immature children: two neonates, two oncology patients, and four transplant recipients. Only two patients initially exhibited evidence of mucocutaneous HSV infection. Blood was cultured for HSV because of perinatal exposure, for routine surveillance, or for the evaluation of fever, esophagitis, or oral lesions in immunocompromised patients. In five cases HSV was recovered only from the blood; in two other instances blood cultures for HSV were the first positive cultures. The time required for the detection of HSV by blood culture ranged from 1 day to 12 days. In one case viremia was transient and cleared without specific therapy. The other seven cases were treated with intravenous acyclovir; in four of these cases, therapy was initiated because of the positive blood culture. The detection of HSV in blood may promote early initiation of antiviral therapy and thereby improve prognosis.

Timely culture for mycobacteria which utilizes a microcolony method.

For the isolation of mycobacteria from clinical specimens, we evaluated a method that used a thinly poured Middlebrook 7H11 agar plate (10 by 90 mm) that was examined microscopically. Inoculated plates were sealed, incubated, and examined at regular intervals for the appearance of microcolonies. Plates were examined microscopically, while still sealed, by focusing on the agar surface through the bottom of the plate and the agar. Plates were scanned at low power (x40 total magnification), and colony morphology was confirmed at intermediate power (x100 to x180 magnification). This method was compared with a traditional method that used macroscopic examination of standard mycobacterial media. By using all specimens submitted for mycobacterial culture over the duration of the study, the method was evaluated until 270 isolates of mycobacteria (Mycobacterium tuberculosis, n = 103; M. avium-M. intracellulare, n = 115; miscellaneous, n = 52) were detected. While the conventional method required an average of 23 days to the time of first detection of mycobacteria, the experimental method required an average of only 11 days. When limited to acid-fast stain-positive specimens that were culture positive for M. tuberculosis, the average interval to positivity was 7 days for the microcolony method compared with 17 days for the conventional method. With the experimental method, the microscopic colonial morphology allowed for the presumptive identification of M. tuberculosis colonies, which were distinguished by cording, and M. avium-M. intracellulare colonies, which were smooth and entire. Presumptive identification was complete for 83.5% of the M. tuberculosis isolates within 10 days and for 85% of the M. avium-M. intracellulare isolates within 11 days after inoculation. If the microcolony method was combined with a conventional tube medium, the composite would optimize for speed of recovery while providing the full sensitivity of the conventional method. In addition to reducing the interval to positivity, the microcolony method allows for the easy detection of mixed mycobacterial infections and yields a presumptive identification that facilitates the selection of a confirmatory gene probe test.

Screening asymptomatic adolescent males for chlamydia.

Ninety-seven asymptomatic 16-21-year-old sexually active adolescent males were evaluated for gonorrhea and chlamydia by culture, chlamydia enzyme immunoassay, and an analysis of a random urine sample for pyuria using centrifuged urine and urine cytometer. The incidence of gonorrhea was 5.3% and chlamydia by culture 12.3%. Immunoassay was superior in sensitivity and specificity (75% and 99%, respectively) to centrifuged urine (sensitivity 58%, specificity 92%) or urine cytometer (58% and 91%) in identifying asymptomatic chlamydia urethritis. Chlamydia enzyme immunoassay is an acceptable, more rapid, and less expensive alternative to culture. The absence of pyuria in asymptomatic males cannot be assumed to indicate the absence of a sexually transmitted disease.

Fever and petechiae in children.

A prospective study of patients with fever and petechiae was performed. Of 190 patients enrolled in the 1-year study, 13 (7%) had meningococcal disease. The most common bacterial association was Streptococcus pyogenes (19 patients). Viral infections were documented in 28 patients. Patients with invasive bacterial disease (group I) appeared more sick, were more likely to have signs of meningeal irritation, and were more likely to have petechiae on the lower extremities than those with less serious, nonbacteremic disease (group II). No patient in group I had petechiae only above the nipple line. Patients in group I had a significantly higher peripheral white blood cell count and absolute band form count. Although no laboratory test or physical finding was sufficiently sensitive to detect all patients with serious disease, the patient with abnormal cerebrospinal fluid, elevated white blood cell count, or elevated absolute band form count was at increased risk for invasive, bacterial disease. Conversely, the risk of serious disease was small if all of these values were in the normal range in the nonill-appearing child or if sore throat and clinical pharyngitis were present in the patient older than 3 years of age.

Comparison of seven kits for detection of rotavirus in fecal specimens with a sensitive, specific enzyme immunoassay.

A sensitive, specific enzyme immunoassay (SSEIA) was compared to four commercial, enzyme-linked immunosorbent assay (ELISA) kits and three latex agglutination assay (LAA) kits: (1) Rotavirus EIA, International Diagnostic Laboratories (IDL), (2) Pathfinder, Kallestadt (KAL), (3) Rotavirus Bio-EnzaBead, Litton (LIT), (4) Rotazyme II, Abbott (RTZII), (5) Slidex Rota-Kit, bioMerieux (SRK), (6) Meritec-Rotavirus, Meridian (MER), and (7) Rotalex, Medical Technology Corporation (RLX). The SSEIA was chosen as the reference method due to its greater sensitivity in comparison to immunoelectron microscopy and polyacrylamide gel electrophoreses of viral RNA segments. Upon evaluation of 136 specimens (of which 44 were positive by SSEIA), the ELISA kits (LIT, KAL, IDL, and RTZII) had sensitivities of 80%, 98%, 91% and 84%; specificities of 95%, 78%, 100%, and 88%; positive predictive values (PPV) of 88%, 68%, 100%, and 77%; and negative predictive values (NPV) of 91%, 99%, 96%, and 92%. When compared with SSEIA, the three LAA tests (SRK, MER, and RLX) had sensitivities of 73%, 75%, and 62%; specificities of 99%, 93%, and 95%; PPVs of 97%, 85%, and 84%; and NPVs of 88%, 89%, and 84%. LAA test results appeared to be reliable, if positive, but the sensitivities of these tests were less than those of the ELISA tests. The ELISA tests that employed specimen specific negative controls were superior in minimizing false positive reactions.

Relation of the outcome of conjunctivitis and the conjunctivitis-otitis syndrome to identifiable risk factors and oral antimicrobial therapy.

The epidemiology, microbiology and clinical outcome of the conjunctivitis-otitis syndrome (CJ-AOM) was investigated in a rural private practice concurrent to a double blind placebo-controlled study of orally administered amoxicillin for prevention of acute otitis media (AOM) secondary to conjunctivitis. Bacterial pathogens were isolated when greater than 15 polymorphonuclear leukocytes/high power field were observed on Gram-stained smear of conjunctival secretions. Nontypable Haemophilus influenzae biotype 2 predominated in CJ-AOM; however, Streptococcus pneumoniae was isolated nearly as frequently as H. influenzae in conjunctivitis without AOM. Younger age (P = 0.001) and more episodes of AOM in the previous year (P = 0.006) were risk factors for CJ-AOM. Persistence of AOM was frequently observed in CJ-AOM. The frequency of AOM secondary to conjunctivitis was reduced (P = .01) in amoxicillin recipients (2 of 41) compared with placebo (11 of 42), but amoxicillin failed to eradicate nasopharyngeal carriage of H. influenzae. More episodes of AOM per year (P less than 0.001) and day care (P less than 0.001) were found to be risk factors for AOM secondary to conjunctivitis.