Optimising the antibiotic treatment of uncomplicated acute appendicitis: a protocol for a multicentre randomised clinical trial (APPAC II trial).

BACKGROUND: Based on epidemiological and clinical data acute appendicitis can present either as uncomplicated (70-80%) or complicated (20-30%) disease. Recent studies have shown that antibiotic therapy is both safe and cost-effective for a CT-scan confirmed uncomplicated acute appendicitis. However, based on the study protocols to ensure patient safety, these randomised studies used mainly broad-spectrum intravenous antibiotics requiring additional hospital resources and prolonged hospital stay. As we now know that antibiotic therapy for uncomplicated acute appendicitis is feasible and safe, further studies evaluating optimisation of the antibiotic treatment regarding both antibiotic spectrum and shorter hospital stay are needed to evaluate antibiotics as the first-line treatment for uncomplicated acute appendicitis. METHODS: APPAC II trial is a multicentre, open-label, non-inferiority randomised controlled trial comparing per oral (p.o.) antibiotic monotherapy with intravenous (i.v.) antibiotic therapy followed by p.o. antibiotics in the treatment of CT-scan confirmed uncomplicated acute appendicitis. Adult patients with CT-scan diagnosed uncomplicated acute appendicitis will be enrolled in nine Finnish hospitals. The intended sample size is 552 patients. Primary endpoint is the success of the randomised treatment, defined as resolution of acute appendicitis resulting in discharge from the hospital without the need for surgical intervention and no recurrent appendicitis during one-year follow-up. Secondary endpoints include post-intervention complications, late recurrence of acute appendicitis after one year, duration of hospital stay, pain, quality of life, sick leave and treatment costs. Primary endpoint will be evaluated in two stages: point estimates with 95% confidence interval (CI) will be calculated for both groups and proportion difference between groups with 95% CI will be calculated and evaluated based on 6 percentage point non-inferiority margin. DISCUSSION: To our knowledge, APPAC II trial is the first randomised controlled trial comparing per oral antibiotic monotherapy with intravenous antibiotic therapy continued by per oral antibiotics in the treatment of uncomplicated acute appendicitis. The APPAC II trial aims to add clinical evidence on the debated role of antibiotics as the first-line treatment for a CT-confirmed uncomplicated acute appendicitis as well as to optimise the non-operative treatment for uncomplicated acute appendicitis. TRIAL REGISTRATION: Clinicaltrials.gov , NCT03236961, retrospectively registered on the 2nd of August 2017.

Iron in boreal river catchments: Biogeochemical, ecological and management implications.

Iron (Fe) is an important element in aquatic ecosystems worldwide because it is intimately tied with multiple abiotic and biotic phenomena. Here, we give a survey of manifold influences of Fe, and the key factors affecting it in the boreal catchments and their waters. It includes the perspectives of biogeochemistry, hydrology, ecology, and river basin management. We emphasize views on the dynamics and impacts of different forms of Fe in riverine environments, including organic colloids and particles, as well as inorganic fractions. We also provide perspectives for land use management in boreal catchments and suggest guidelines for decision making and water management. Based on our survey, the main emphases of water protection and management programs should be (i) prevention of Fe mobilization from soil layers by avoiding unnecessary land-use activities and minimizing soil disturbance in high-risk areas; (ii) disconnecting Fe-rich ground water discharge from directly reaching watercourses; and (iii) decreasing transport of Fe to watercourses by applying efficient water pollution control approaches. These approaches may require specific methods that should be given attention depending on catchment conditions in different areas. Finally, we highlight issues requiring additional research on boreal catchments. A key issue is to increase our understanding of the role of Fe in the utilization of DOM in riverine food webs, which are typically highly heterotrophic. More knowledge is needed on the metabolic and behavioral resistance mechanisms that aquatic organisms, such as algae, invertebrates, and fish, have developed to counter the harmful impacts of Fe in rivers with naturally high Fe and DOM concentrations. It is also emphasized that to fulfil the needs presented above, as well as to develop effective methods for decreasing the harmful impacts of Fe in water management, the biogeochemical processes contributing to Fe transport from catchments via rivers to estuaries should be better understood.

Extremely high prevalence of multidrug resistant tuberculosis in Murmansk, Russia: a population-based study.

Drug resistance and molecular epidemiology of tuberculosis (TB) in the Murmansk region was investigated in a 2-year, population-based surveillance of the civilian population. During 2003 and 2004, isolates from all culture-positive cases were collected (n = 1,226). Prevalence of multi-drug resistance (MDR) was extremely high, as 114 out of 439 new cases (26.0%), and 574 out of 787 previously treated cases (72.9%) were resistant to at least isoniazid (INH) and rifampin (RIF). Spoligotyping of the primary MDR-TB isolates revealed that most isolates grouped to the Beijing SIT1 genotype (n = 91, 79.8%). Isolates of this genotype were further analyzed by IS6110 RFLP. Sequencing of gene targets associated with INH and RIF resistance further showed that the MDR-TB strains are highly homogeneous as 78% of the MDR, SIT1 strains had the same resistance-conferring mutations. The genetic homogeneity of the MDR-TB strains indicates that they are actively transmitted in Murmansk.

IL-6 and other biomarkers as predictors of severity in COVID-19.

OBJECTIVE: Cytokine release syndrome is suggested to be the most important mechanism triggering acute respiratory distress syndrome and end organ damage in COVID-19. The severity of disease may be measured by different biomarkers. METHODS: We studied markers of inflammation and coagulation as recorded in 29 patients on admission to the hospital in order to identify markers of severe COVID-19 and need of ICU. RESULTS: Patients who were eventually admitted to ICU displayed significantly higher serum levels of interleukin-6 (IL-6), C-reactive protein (CRP), and procalcitonin. No statistical differences were found between the groups in median levels of lymphocytes, D-dimer or ferritin. CONCLUSIONS: IL-6 and CRP were the strongest predictors of severity in hospitalized patients with COVID-19.

Prospective evaluation of pyrosequencing for the rapid detection of isoniazid and rifampin resistance in clinical Mycobacterium tuberculosis isolates.

A pyrosequencing-based method for the rapid detection of isoniazid (INH) and rifampin (RIF) resistance in Mycobacterium tuberculosis was evaluated in clinical practice. The method can detect the INH resistance-causing katG315 mutation, and all mutations in the RIF resistance-determining rpoB core region, in less than 6 h from cultured isolates. The method was first validated with 42 isolates, and was subsequently prospectively evaluated with 91 isolates, including clinical isolates and external quality control assessment strains, over a period of 2.5 years. The pyrosequencing results of clinical isolates were available, on average, 19 days earlier (median 19 days; range 3-43 days) than conventional susceptibility testing results. The composite data showed that the sensitivity of pyrosequencing for detecting resistance correctly was 66.7% for INH and 97.4% for RIF. The specificity of pyrosequencing was 100% for both drugs. Acceptable sensitivity for detecting resistance and the rapidness of pyrosequencing make it a valuable tool in the clinical setting.

Molecular genetics of drug-resistant Mycobacterium tuberculosis isolates in Finland, 1995-2004.

SETTING: Modern molecular methods help us to understand the transmission and epidemiology of Mycobacterium tuberculosis. OBJECTIVE: To analyse the molecular epidemiology of drug-resistant tuberculosis (TB), and to characterise isoniazid (INH) and rifampicin (RMP) resistance conferring mutations in Finland during 1995-2004. DESIGN: A total of 3959 new M. tuberculosis isolates underwent drug susceptibility testing; all phenotypically resistant isolates were genotyped by IS6110 restriction fragment length polymorphism and spoligotyping if necessary. INH- and/or RMP-resistant isolates were sequenced for their resistance associated genes, katG locus 315 and rpoB, respectively. RESULT: Of the 3959 isolates tested (92.4% of culture-positive cases), 183 (4.6%) were resistant to at least one first-line anti-tuberculosis drug; 14 (0.4%) isolates were multidrug-resistant. Thirty-seven (20.4%) resistant isolates belonged to 17 clusters, and the largest cluster included four isolates. The Beijing family genotype accounted for 8.8% (16 isolates) of all drug-resistant isolates. A Ser315Thr mutation in katG was found in 46.7% (56 isolates) of the INH-resistant isolates and rpoB was mutated in 85.7% (18 isolates) of the isolates resistant to RMP. CONCLUSION: Transmission of drug-resistant TB is rare in Finland, especially between indigenous and immigrant populations. Screening of mutations that confer INH and RMP resistance seems to be feasible if risk factors for multidrug resistance exist.

Evaluation of a novel strip test, GenoType Mycobacterium CM/AS, for species identification of mycobacterial cultures.

A novel DNA strip assay, GenoType Mycobacterium AS, was evaluated for its ability to identify 219 mycobacterial isolates in combination with the GenoType Mycobacterium CM assay. The results were compared with those obtained by conventional 16S rDNA sequencing. The Genotype test correlated well (96%) with sequencing. However, with the CM kit alone, it was possible to identify most (88%) of the isolates found in clinical specimens, and the AS kit provided very little additional information.

Early dissemination of Borrelia burgdorferi without generalized symptoms in patients with erythema migrans.

The diagnosis of erythema migrans (EM) is not always easy, and reports of culture- or PCR-confirmed diagnosis as well as reports of EM with simultaneous disseminated disease are few. Characteristics and incidence of EM in addition to frequency of early dissemination of B. burgdorferi were studied in the archipelago of South-Western Finland prospectively using questionnaires, skin biopsies and blood samples. Clinical EM was recognized in 82 patients (incidence 148/100,000 inhabitants/year). Of skin biopsy samples, 35.5% were positive by PCR (the majority B. garinii), and 21.5% by cultivation (all B. garinii). Of blood samples, 3.8% were positive by PCR, and 7.7% by cultivation. Of the patients, 30.9% were seropositive at the first visit, and 52.9% 3 weeks later. Of the patients with laboratory confirmed diagnosis, the EM lesion was ring-like in 31.8% and homogeneous in 65.9%. Dissemination of B. burgdorferi, based on culture or PCR positivity of blood samples, was detected in 11.0% of the patients. The frequency of generalized symptoms was nearly the same in patients with as in those without dissemination (22.2% vs 27.4%). Only 21.4% of the patients with culture-positive EM recalled a previous tick bite at the site of the EM lesion. We conclude that EM lesions are more often homogeneous than ring-like. B. burgdorferi may disseminate early without generalized symptoms.

Performance of BACTEC 960 Mycobacteria growth indicator tube in the susceptibility testing of genetically characterized Mycobacterium tuberculosis isolates.

In this study, the 7H10 agar proportion method was compared with the BACTEC TB-460 and BACTEC MGIT 960 systems (BD Biosciences, USA) for the susceptibility testing of 22 genetically characterized Mycobacterium tuberculosis isolates for isoniazid, rifampin, streptomycin, and ethambutol. The 7H10 agar proportion method agreed with the resistant genotype in 87.3%, BACTEC TB-460 in 92.7%, and the MGIT in 96.4% of the cases, showing the high sensitivity of MGIT in the detection of resistant isolates.

LCx Mycobacterium tuberculosis assay is valuable with respiratory specimens, but provides little help in the diagnosis of extrapulmonary tuberculosis.

BACKGROUND: Commercial nucleic acid amplification tests, designed for the detection of Mycobacterium tuberculosis DNA/RNA in respiratory samples, are often applied also in nonrespiratory specimens in order to verify the diagnosis of extrapulmonary tuberculosis. AIM. To evaluate the value of the Abbott LCx Mycobacterium tuberculosis assay for the diagnosis of pulmonary and extrapulmonary tuberculosis based on routine clinical laboratory results. METHODS: The assay was used to analyse 350 respiratory and 826 nonrespiratory specimens from 961 patients, of whom 3.6% had culture-proven tuberculosis. The results obtained by the LCx assay were compared with the records on mycobacterial isolates of the national reference laboratory and, in the case of positive findings, with clinical data. RESULTS: In comparison with culture, the sensitivity, specificity and positive/negative predictive value of the assay on respiratory specimens were 87.5%, 99.7%, 93.3% and 99.4%, respectively. With nonrespiratory specimens, the overall sensitivity, specificity and positive/negative predictive value of the LCx assay were 73.3%, 98.0%, 40.7% and 99.5%, respectively. When clinical and histological data were also included, the positive predictive value of LCx with nonrespiratory specimens was 45.8%. CONCLUSION: Critical interpretation of the nucleic acid amplification results obtained from nonrespiratory specimens is necessary in both laboratory and clinical settings.

katG mutations in isoniazid-resistant Mycobacterium tuberculosis isolates recovered from Finnish patients.

katG and inhA genes from isoniazid-resistant Mycobacterium tuberculosis strains isolated in Finland were examined by PCR or sequencing. By PCR, katG was not detected in 3 of 54 strains. Sequencing of katG from 13 strains showed small point mutations or insertions; a previously described mutation causing a Ser-to-Thr change at position 315 was found in 4 strains, and there were nine new missense mutations of katG. A 209-bp segment of inhA from 17 strains was sequenced, but no mutations were observed. This result indicates that different mutations prevail in different geographical areas.

Prospective evaluation of the GenoType assay for routine identification of mycobacteria.

In order to evaluate the proficiency of the GenoType Mycobacteria strip hybridization assay (Hain Lifescience, Nehren, Germany) for the routine identification of mycobacteria, the assay was used to identify 178 clinical isolates during a 6-month prospective study. The GenoType results were compared to the identification results obtained with AccuProbe (GenProbe, San Diego, CA, USA) or 16S rDNA sequencing, and an overall agreement of 89.3% between GenoType and the two reference methods was reached. The GenoType assay is, thus, a rapid and reliable method for the identification of clinically important mycobacteria, and it is well suited for use in a routine laboratory.

Line probe assay in the rapid detection of rifampin-resistant Mycobacterium tuberculosis directly from clinical specimens.

The sensitivity of a PCR-based line probe assay (Inno-LiPA Rif. TB Assay; Innogenetics NV Zwijndrecht, Belgium) was studied by using nested-PCR technique. A total of 75 specimens, representing various body locations from 70 suspected tuberculosis patients were obtained. LiPA yielded 30 Mycobacterium tuberculosis complex positive results (sensitivity 58.8%, compared with final diagnoses) whereas culture for M. tuberculosis was positive in 18 specimens (sensitivity 35.3%). Genotypic rifampin resistance testing by LiPA showed that 7 specimens contained rpoB mutations associated with RMP resistance, and sequencing data of the rpoB gene and LiPA patterns agreed in 29 of 30 M. tuberculosis positive specimens (96.7%). This indicates reliable performance, which makes the test suitable for the rapid determination of resistance to rifampin directly in clinical samples. However, the best results are obtained if LiPA is combined with conventional staining and culture methods.

Rapid detection of rifampin-resistant Mycobacterium tuberculosis by sequencing and line probe assay.

Resistance to rifampin (RMP) is associated with mutations in the rpoB gene. Disk elution method, direct DNA sequencing and line probe assay were compared in rapid detection of rpoB mutations from 30 Mycobacterium tuberculosis isolates. Concordance between LiPA and culture was 93.3%, whereas sequencing yielded 100% concordance with culture. These results indicate that line probe assay is nearly as sensitive a method as sequencing in rapid detection of RMP-resistance of M. tuberculosis isolates, and can be applied in laboratories working with standard PCR equipment.

A Ser315Thr substitution in KatG is predominant in genetically heterogeneous multidrug-resistant Mycobacterium tuberculosis isolates originating from the St. Petersburg area in Russia.

Parts of katG and rpoB from 27 Russian Mycobacterium tuberculosis isolates were sequenced to detect mutations causing resistance to isoniazid (INH) and rifampin (RMP), respectively. All 24 INH-resistant isolates had a mutated katG, and 22 of them (91.7%) carried a mutation coding for a Ser315Thr shift. An rpoB mutation was noted for each of the 21 RMP-resistant isolates, with Ser531Leu being the most prevalent change encoded. Only two isolates had identical IS6110 fingerprints.

pncA mutations in pyrazinamide-resistant Mycobacterium tuberculosis isolates from northwestern Russia.

Thirty-six pyrazinamide-resistant and eight pyrazinamide-susceptible Mycobacterium tuberculosis isolates from Russia were analyzed for their pncA mutations. Thirty-one (86.1%) of the resistant isolates had a mutation either in pncA or upstream of the gene. Twenty of the 23 different mutations found in this study had not been described earlier. pncA genotype correlated well with pyrazinamidase activity and BACTEC 460 susceptibility test results.