The Spanish Society of Epidemiology and Oral Public Health.

In 2019 the Sociedad Española de Epidemiología y Salud Pública Oral (SESPO, The Spanish Society of Epidemi- ology and Oral Public Health) approached BASCD and EADPH about using Community Dental Health as their offi cial journal. Very pleasant discussions between all parties mean that henceforth, SESPO will translate the abstracts of all papers published in Community Dental Health so that they may be placed on our website. The SESPO website will link to the translated abstracts.

GenoType MTBDRsl for molecular detection of second-line-drug and ethambutol resistance in Mycobacterium tuberculosis strains and clinical samples.

The purpose of this study was to evaluate the GenoType MTBDRsl assay (Hain Lifescience GmbH, Nehren, Germany) for its ability to detect resistance to fluoroquinolones (FLQ), injectable second-line antibiotics [kanamycin (KM) and capreomycin (CM)], and ethambutol (EMB) in Mycobacterium tuberculosis clinical strains and directly in clinical samples. A total of 34 clinical strains were characterized with the Bactec 460 TB system. Fifty-four clinical samples from 16 patients (5 were smear negative and 49 were smear positive) were also tested directly. The corresponding isolates of the clinical specimens were also analyzed with the Bactec 460TB. When there was a discrepancy between assays, pyrosequencing was performed. The overall rates of concordance of the MTBDRsl and the Bactec 460TB for the detection of FLQ, KM/CM, and EMB susceptibility in clinical strains were 72.4% (21/29), 88.8% (24/27), and 67.6% (23/34), whereas for clinical samples, rates were 86.5% (45/52), 92.3% (48/52), and 56% (28/50), respectively. In conclusion, the GenoType MTBDRsl assay may be a useful tool for making early decisions regarding KM/CM susceptibility and to a lesser extent regarding FLQ and EMB susceptibility. The test is able to detect mutations in both clinical strains and samples with a short turnaround time. However, for correct management of patients with extensively drug-resistant tuberculosis, results must be confirmed by a phenotypical method.

Usefulness of consecutive biomarkers measurement in the management of community-acquired pneumonia.

The aim of this study was to investigate whether procalcitonin (PCT), neopterin, C-reactive protein (CRP), and mid regional pro-atrial natriuretic peptide (MR-proANP) levels at admission and during the clinical course can be useful for the management of patients with pneumonia. The study population consisted of 75 patients with clinical and radiological diagnosis of pneumonia. Serum samples were collected at admission and during hospitalization. Complications were defined as intensive care unit (ICU) admission or death. The levels of PCT were significantly higher in pneumonia of definite bacterial origin in comparison to probable bacterial or unknown origin. The PCT levels were higher in pneumococcal pneumonia. The PCT and MR-proANP levels increased significantly according to the Pneumonia Severity Index (PSI). All biomarkers levels are higher in patients developing complications and who were dying. The serial levels of MR-proANP remain significantly elevated in patients developing complications and in patients classified in PSI and CURB-65 risk groups. In patients not developing complications, there is a significant decrease in the PCT levels. PCT can be useful for identifying pneumonia etiology. PCT and MR-proANP levels correlate with pneumonia severity rules. PCT and MR-proANP serial measurements can be useful for predicting short-term prognosis. Systemic biomarkers can provide additional information regarding clinical evolution, because these are dynamic and can be measured daily.

Diagnosing TB infection in children: analysis of discordances using in vitro tests and the tuberculin skin test.

The aim of the present study was to compare the performance of the interferon (IFN)-γ tests (QuantiFERON®-TB Gold In-Tube (QFT-G-IT) and T-SPOT®.TB) with the tuberculin skin test (TST) in diagnosing tuberculosis (TB) infection in children, and to analyse discordant results. This was a prospective study including 98 children from contact-tracing studies and 68 children with TST indurations ≥ 5 mm recruited during public health screenings. Positive IFN-γ tests results were associated with risk of exposure (p<0.0001). T-SPOT.TB was positive in 11 (78.6%) out of 14 cases with active TB and QFT-G-IT in nine (64.3%) out of 14 cases. Sensitised T-cells against Mycobacterium avium were detected in six out of 12 children not vaccinated with bacille Calmette-Guérin (BCG), a TST induration 5-9 mm in diameter and both IFN-γ tests negative. In concordant IFN-γ tests results, a positive correlation was found (p = 0.0001) between the number of responding cells and the amount of IFN-γ released. However, in discordant IFN-γ tests results this correlation was negative (p = 0.371): an increase in the number of spot-forming cells correlated with a decrease in the amount of IFN-γ released. The use of IFN-γ tests is helpful for the diagnosis of TB infection, avoiding cross-reactions with BCG immunisation and nontuberculous mycobacterial infections. The analysis of highly discordant results requires further investigation to elucidate possible clinical implications.

Pyrosequencing for rapid molecular detection of rifampin and isoniazid resistance in Mycobacterium tuberculosis strains and clinical specimens.

The aim of this study was to evaluate a pyrosequencing method for the detection of Mycobacterium tuberculosis isolates resistant to rifampin and isoniazid using both clinical strains and clinical samples, comparing the results with those of the Bactec 460TB and GenoType MTBDRplus assays. In comparison to Bactec 460TB as the gold standard, the sensitivity of pyrosequencing for detecting isoniazid and rifampin resistance was 76.9% and 97.2%, respectively, for clinical strains, and the specificity was 97.2 and 97.9%, respectively. For clinical specimens, the sensitivity and specificity for both drugs were 85.7% and 100%, respectively. The overall concordance between pyrosequencing and the GenoType MTBDRplus assay for clinical strains was 99.1%, and for clinical samples, it was 98.2%. Pyrosequencing is a valuable tool for rifampin and isoniazid resistance detection.

Evaluating the non-tuberculous mycobacteria effect in the tuberculosis infection diagnosis.

The aim of the present study was to determine the role of previous non-tuberculous mycobacteria sensitisation in children as a factor of discordant results between tuberculin skin test (TST) and an in vitro T-cell based assay (T-SPOT.TB; Oxford Immunotec, Oxford, UK). We enrolled 21 non-bacille Calmette-Guérin-vaccinated paediatric patients for suspicious of latent tuberculosis infection (LTBI). These patients yielded a positive TST and a negative T-SPOT.TB. Cells were stimulated with Mycobacterium avium sensitin (having cross-reaction with Mycobacterium intracellulare and Mycobacterium scrofulaceum) and the presence of reactive T-cells was determined by an ex vivo ELISPOT. From the 21 patients, in 10 cases (47.6%), we obtained a positive ELISPOT result after stimulation with M. avium sensitin, in six (28.6%) cases, the result was negative and in the remaining five (23.8%) cases, the result was indeterminate. In conclusion, previous non-tuberculous mycobacteria sensitisation induces false-positive results in the TST for diagnosing LTBI and the use of gamma-interferon tests could avoid unnecessary chemoprophylaxis treatment among a child population.

The tuberculin skin test increases the responses measured by T cell interferon-gamma release assays.

RUTI is a vaccine consisting of Mycobacterium tuberculosis bacilli grown in stress conditions that is fragmented, detoxified and liposomed. RUTI was designed to shorten the treatment of latent tuberculosis infection (LTBI) with isoniazid from 9 months to just 1 month, by additional treatment with two inoculations of RUTI 4 weeks apart. During the validation process for monitoring the immunogenicity of administration of RUTI in a Phase I clinical trial, the question arose whether to introduce the tuberculin skin test (TST) in the screening of non-LTBI volunteers. This study was designed to evaluate the effect of TST on subsequent different T-cell interferon-gamma release assay (TIGRA) responses, using a spectrum of M. tuberculosis-related antigens (ESAT-6, CFP-10, 16 kDa, 19 kDa, MPT64, Ag 85B, 38 kDa, hsp65, PPD and BCG). The results showed an increase in post-TST response even in non-LTBI subjects for most antigens tested, as measured both by whole blood assay (WBA) and ELISPOT. Increased ELISPOT response decreased toward pre-TST levels within 1 month whereas the WBA response did not. Taking into account that there is no definitive correlation between TST and TIGRA tests to diagnose LTBI and the feasibility that TST might alter the immune monitoring included in clinical trials, these data suggest that TST determination should be carefully planned to avoid any interference with TIGRA.

Mycobactericidal activity of chlorine dioxide wipes in a modified prEN 14563 test.

Tristel Sporicidal Wipes are chlorine dioxide-based disinfectant wipes for disinfecting non-lumened semi-critical medical devices. In this study, the mycobactericidal activity of this product was assessed by a modified version of the European Standard prEN 14563 carrier test under clean conditions against Mycobacterium avium. The chlorine dioxide concentration in the activated wipe was 200ppm. The results showed that the chlorine dioxide wipes were mycobactericidal in 30s contact time with mechanical action and in 60s without mechanical action, both under clean conditions.

Evaluation of a Legionella urinary antigen enzyme immunoassay for rapid detection of Legionella pneumophila in water samples.

Despite advances in medium formulations and pretreatment techniques, recovery of Legionella from water samples can still be quite low, difficult and time consuming. The aim of this study was to evaluate the utility of a Legionella urinary antigen enzyme immunoassay (Bartels ELISA, Trinity Biotech, Ireland) for the detection of Legionella in water samples. Reference ATCC Legionella strains were used to spike water samples to a final concentration of 10(4)-10(5)cfu/ml. The lower detection limit of the test for all Legionella pneumophila serogroups was assessed by serial dilutions of spiked water samples. Legionella antigen was detected in all filtered samples except for those spiked with L. bozemanii and L. longbeachae. The lower detection limit for soluble L. pneumophila serogroup 1 antigen was 780cfu/ml. Bartels ELISA could be a useful method for antigen detection in water samples when a high recovery of L. pneumophila is suspected. The test could be used as a rapid screening method for the detection of Legionella in a large number of samples. However, the low sensitivity of the test requires to keep on performing conventional culture for isolation and for further studies on isolated bacteria.

Investigating intracellular persistence of Staphylococcus aureus within a murine alveolar macrophage cell line.

OBJECTIVE: Staphylococcus aureus is a particularly difficult pathogen to eradicate from the respiratory tract. Previous studies have highlighted the intracellular capacity of S.aureus in several phagocytic and non-phagocytic cells. The aim of this study was to define S.aureus interaction within a murine alveolar macrophage cell line. METHODS: Cell line MH-S was infected with Newman strain. Molecular mechanisms involved in phagocytosis were explored. To assess whether S.aureus survives intracellularly quantitative (gentamicin protection assays and bacterial plating) and qualitative analysis (immunofluorescence microscopy) were performed. Bacterial colocalization with different markers of the endocytic pathway was examined to characterize its intracellular trafficking. RESULTS: We found that S.aureus uptake requires host actin polymerization, microtubule assembly and activation of phosphatidylinositol 3-kinase signaling. Time course experiments showed that Newman strain was able to persist within macrophages at least until 28.5 h post infection. We observed that intracellular bacteria are located inside an acidic subcellular compartment, which co-localizes with the late endosome/lysosome markers Lamp-1, Rab7 and RILP. Colocalization counts with TMR-dextran might reflect a balance between bacterial killing and intracellular survival. CONCLUSIONS: This study indicates that S.aureus persists and replicates inside murine alveolar macrophages, representing a privileged niche that can potentially offer protection from antimicrobial activity and immunological host defense mechanisms.

Comparison of a monoclonal with a polyclonal antibody-based enzyme immunoassay stool test in diagnosing Helicobacter pylori infection before and after eradication therapy.

BACKGROUND: Detection of Helicobacter pylori antigen in stool samples has been a subject of controversy. However, it has been included in several clinical guidelines as a recommended non-invasive testing procedure in dyspeptic patients. AIM: To compare a monoclonal enzyme immunoassay for detection of H. pylori stool antigen (Amplified IDEIA HpStAR, DakoCytomation) with a polyclonal enzyme immunoassay (HpSA test, Premier Platinum HpSA, Meridian Diagnostics) in diagnosing infection and in determining H. pylori status after eradication treatment. METHODS: We evaluated stool samples of 198 patients diagnosed with H. pylori infection and of 41 patients without infection. The results of the monoclonal enzyme immunoassay HpStAR were compared with those of the polyclonal enzyme immunoassay HpSA. RESULTS: The sensitivity and specificity of HpStAR were 91.9% and 70.7%, while those of HpSA were 89.4% and 80.5%, respectively. In the 126 patients evaluated 6 weeks after eradication therapy, the overall agreement between urea breath test and HpStAR was 90.5% (P = 0.710) and between urea breath test and HpSA was 76.9% (P = 0.410). CONCLUSIONS: HpStAR is a rapid and easy-to-perform test with similar sensitivity to HpSA in the diagnosis of H. pylori infection, although it had lower specificity. In contrast, HpStAR is more accurate after eradication therapy than HpSA.

Usefulness of pneumococcal antigen detection in pleural fluid samples by immunochromatographic assay for diagnosis of pneumococcal pneumonia.

This study investigated the utility of an immunochromatographic test (ICT) for the detection of Streptococcus pneumoniae antigens in pleural fluid. Antigen was detected in 15 of 19 (79%) patients with pneumococcal pneumonia. The ICT was always negative in patients with non-pneumococcal pneumonia, but was positive in three cases with a non-infectious aetiology. In patients with pneumonia for which no pathogen was identified, antigen was detected in one of 24 pleural fluids tested. The ICT can be a valuable tool for the management of pneumonia because it can detect pneumococcal antigen in pleural effusion samples.

Mycobactericidal and tuberculocidal activity of Korsolex AF, an amine detergent/disinfectant product.

The mycobactericidal and tuberculocidal activities of Korsolex AF against Mycobacterium tuberculosis, Mycobacterium avium-Mycobacterium intracellulare (MAI), Mycobacterium kansasii and Mycobacterium chelonae were determined using quantitative suspension and carrier tests. The effects of organic load and hard water were also considered. A clinical isolate of MAI was the most resistant of the four test organisms. A 2% solution had good mycobactericidal and tuberculocidal activities after 30 min of exposure. Although further evaluation using European standard tests is necessary, we conclude that Korsolex AF appears to be a promising product for the disinfection of hospital instruments contaminated with mycobacteria.

[Variability in Epstein-Barr virus serological markers in adult kidney transplant recipients]. / Variabilidad en los marcadores serológicos del virus de Epstein-Barr en receptores adultos de trasplante renal.

Epstein-Barr virus (EBV) infection is associated with the development of post-transplant lymphoproliferative disorders (PTLD). However, the clinical relevance and criteria for EBV serological reactivation in EBV-seropositive transplant recipients is unclear. EBV-specific antibodies: viral capsid immunoglobulm G [IgG (VCA)], nuclear antigen (EBNA) IgG, immunoglobulin M [IgM (VCA)] and early antigen IgG (EA) were prospectively analyzed in 71 adult kidney transplant recipients, before starting immunosuppression, when they were uraemic, and after transplantation. A total of 351 serum samples were tested. Relevance of different EBV reactivation-related variables were analyzed using the chi-square test. In 37 of 71 (52.1%) patients IgM (VCA) or IgG (EA) were detected when they were uraemic. EBV reactivation occurred in 25 of 71 (35.2%) patients, with clinical symptoms (fever, leukopenia, kidney function impairment, and increase in transaminases) in nine cases. One of 71 patients developed a PTLD, without detection of serologically EBV reactivation, but with an increase in EBV viral load. Absence of mycophenolate mofetil, that inhibits lymphocyte proliferation and antibody production, in immunosuppression was statistically significantly associated with EBV reactivation (p = 0.015). Serological diagnosis of EBV reactivation should be based on strict criteria (IgM (VCA) seroconversion, four-fold increase in IgM (VCA) or IgG (EA), or four-fold decrease in IgG (EBNA) titers and on analysis of serial samples. Some EBV-seropositive patients at high risk of developing PTLD could benefit from this diagnostic methodology.

AID TB resistance line probe assay for rapid detection of resistant Mycobacterium tuberculosis in clinical samples.

OBJECTIVES: To determine the sensitivity and specificity of AID TB Resistance line probe assay (AID Diagnostika, Germany) to detect Mycobacterium tuberculosis and its resistance to first- and second-line drugs in clinical samples using BACTEC 460TB as the reference standard. METHODS: The test consists on three strips to detect resistance to isoniazid/rifampicin, fluoroquinolones/ethambutol, and kanamycin/amikacin/capreomycin/streptomycin, respectively. This test was performed on 65 retrospectively selected clinical samples corresponding to 32 patients. RESULTS: A valid result was obtained for 92.3% (60/65), 90.8% (59/65) and 78.5% (51/65) of the samples tested, considering the three strips, respectively. Global concordance rates between AID and BACTEC for detecting resistance to isoniazid, rifampicin, fluoroquinolones, ethambutol, kanamycin/capreomycin and streptomycin were 98.3% (59/60), 100% (60/60), 91.5% (54/59), 72.9% (43/59), 100% (51/51) and 98.0% (50/51), respectively. Regarding the discordant results obtained between AID and BACTEC, the alternative molecular methods performed (GenoType MTBDRplus, GenoType MTBDRsl [Hain Lifescience, Germany] and/or pyrosequencing) confirmed the genotypic result in 90.9% (20/22) of the cases. CONCLUSIONS: AID line probe assay is a useful tool for the rapid detection of drug resistance in clinical samples enabling an initial therapeutic approach. Nevertheless, for a correct management of drug resistant tuberculosis patients, molecular results should be confirmed by a phenotypic method.

Diagnostic accuracy study of multiplex PCR for detecting tuberculosis drug resistance.

OBJECTIVE: To study the diagnostic accuracy of a multiplex real-time PCR (Anyplex II MTB/MDR/XDR, Seegene, Corea) that detects Mycobacterium tuberculosis resistant to isoniazid (INH), rifampicin (RIF), fluoroquinolones (FLQ) and injectable drugs (kanamycin [KAN], amikacin [AMK] and capreomycin [CAP]) in isolates and specimens. METHODS: One hundred fourteen cultured isolates and 73 sputum specimens were retrospectively selected. Results obtained with multiplex PCR were compared with those obtained with BACTEC. Discordant results between multiplex PCR and BACTEC were tested by alternative molecular methods. RESULTS: Sensitivity and specificity of multiplex PCR for detecting drug resistance in isolates were 76.5% and 100%, respectively, for INH; 97.2% and 96.0%, respectively, for RIF; 70.4% and 87.9%, respectively, for FLQ; 81.5% and 84.8%, respectively, for KAN; 100% and 60%, respectively, for AMK, and 100% and 72.3%, respectively, for CAP. Sensitivity and specificity of Anyplex for detecting drug resistance in specimens were 93.3% and 100%, respectively, for INH; 100% and 100%, respectively, for RIF; 50.0% and 100%, respectively, for FLQ; and 100% and 94.4%, respectively, for both KAN and CAP. Among the discordant results, 87.7% (71/81) of results obtained with the multiplex PCR were concordant with at least one of the alternative molecular methods. CONCLUSIONS: This multiplex PCR may be a useful tool for the rapid identification of drug resistant tuberculosis in isolates and specimens, thus allowing an initial therapeutic approach. Nevertheless, for a correct management of patients, results should be confirmed by a phenotypic method.

Distribution of surface-exposed antigenic glycolipids in recent clinical isolates of Mycobacterium tuberculosis.

The distribution of surface-exposed antigenic glycolipids in seven recent clinical isolates of Mycobacterium tuberculosis was established. Thin-layer and liquid chromatographies revealed a uniformity in the glycolipid pattern. Chemical analysis of the individual glycolipids of a selected strain enabled the identification of glycolipids of serological interest in all the other clinical isolates. Phenolic glycolipid-Tb1 (PGL-Tb1) was lacking in all strains, but appreciable amounts of a partially deglycosylated version (PGL-Tb1D) were present in the seven isolates. Diacyltrehaloses (DATs) were detected in all strains, showing themselves to be major glycolipids. Lipooligosaccharides (LOS-II) were present in the seven strains studied though only in trace amounts. These results shed new light on the open debate on the distribution of these interesting glycolipids in typical clinical isolates of M. tuberculosis. In the search for a serological test for tuberculosis, and in accordance with our observations, we believe that PGL-Tb1 and LOS-II should not be the target molecules for serology and that it is worthwhile to continue investigating the value of DATs as antigens. We also believe that it would be of interest to undertake research to assess the usefulness of PGL-Tb1D as an antigen.

Impact of previous antimicrobial therapy on the etiology and outcome of ventilator-associated pneumonia.

OBJECTIVE: To define the influence of prior antibiotic use on the etiology and mortality of ventilator-associated pneumonia (VAP). SETTING: A university hospital medical-surgical ICU. DESIGN: Prospective clinical study. METHODS: Over a 35-month period, we prospectively studied 129 consecutive episodes of VAP. Etiologic diagnosis was established using a protected specimen brush and quantitative culture techniques. We examined prognostic factors by univariate and multivariate analyses using a statistical software package (SPSS). RESULTS: The rate of VAP caused by Gram-positive cocci or Haemophilus influenzae was statistically lower (p < 0.05) in the patients who had received antibiotics previously, while the rate of VAP caused by Pseudomonas aeruginosa was statistically higher (p < 0.01). Patients died of causes directly related to the infection in 18 (14.0 percent) episodes, P aeruginosa being isolated in 9 of these fatal cases. Indeed, we found that 27.7 percent (15/54) of patients who had received prior antimicrobial therapy before the onset of pneumonia died, compared with only 4.0 percent (3/75) of those who did not. In the univariate analysis, the variables significantly associated with attributable mortality were age older than 45 years, use of corticosteroids, presence of shock, hospital day of VAP over 9, antecedent COPD, and a prior antibiotic use. A step-forward logistic regression analysis defined only prior antibiotic use (p < 0.0001, OR = 9.2) as significantly influencing the risk of death from VAP. The same result was obtained when severity was included in the model. However, prior antibiotic use entirely dropped out as a significant risk factor when the etiologic agent was included in the regression equation. CONCLUSIONS: Distribution of infecting microorganisms responsible for VAP differs in patients who received prior antimicrobial therapy, and this factor determines a higher mortality rate. We suggest a restrictive antibiotic policy in mechanically ventilated patients with the purpose of reducing the risk of death from VAP.

A three-year study of severe community-acquired pneumonia with emphasis on outcome.

Fifty-eight consecutive patients with severe community-acquired pneumonia were studied prospectively during a three-year period. The group included 44 men and 14 women (mean age: 45.0 +/- 15.7 years). The cause of pneumonia was diagnosed in 35 (60.3 percent) cases, and the most common pathogens were Streptococcus pneumoniae (37.1 percent), Legionella pneumophila (22.8 percent) and Gram-negative bacilli (11.4 percent). The fact that Mycobacterium tuberculosis was present in four (11.4 percent) patients and Pneumocystis carinii in three (8.5 percent) is worthy of note. The overall death rate was 22.4 percent. More than 50 percent of deaths occurred within the first five days and were caused by septic shock, hemoptysis (tuberculosis) or hypoxia. However, hypoxia remains the main fatal complication and all late-occurring deaths (> 5 days) observed were due to this cause. These data could be important in planning strategies and protocols to improve prognosis.

Pneumonia due to Haemophilus influenzae among mechanically ventilated patients. Incidence, outcome, and risk factors.

Incidence and potential risk factors for pneumonia due to Haemophilus influenzae in adults treated with mechanical ventilation in a medical-surgical ICU were investigated. Diagnosis was established in 91 episodes and H influenzae was isolated in 20 of them. Mean onset of ventilator-associated pneumonia (VAP) due to H influenzae was 10.8 days after intubation. Six patients with H influenzae VAP died in the ICU. Of 13 risk factors for developing VAP due to H influenzae, an absence of prior antibiotic treatment was the only variable which had statistical significance (p < 0.001). In these mechanically ventilated patients, Haemophilus influenzae was a common causative agent for VAP, frequently associated with Gram-positive cocci. Episodes of H influenzae VAP were associated with a lower mortality compared with other etiologies. The epidemiologic and clinical findings indicate that patients without a prior antimicrobial treatment have increased susceptibility to infections of the airway by H influenzae.