Country-report pattern corrections of new cases allow accurate 2-week predictions of COVID-19 evolution with the Gompertz model.

Accurate short-term predictions of COVID-19 cases with empirical models allow Health Officials to prepare for hospital contingencies in a two-three week window given the delay between case reporting and the admission of patients in a hospital. We investigate the ability of Gompertz-type empiric models to provide accurate prediction up to two and three weeks to give a large window of preparation in case of a surge in virus transmission. We investigate the stability of the prediction and its accuracy using bi-weekly predictions during the last trimester of 2020 and 2021. Using data from 2020, we show that understanding and correcting for the daily reporting structure of cases in the different countries is key to accomplish accurate predictions. Furthermore, we found that filtering out predictions that are highly unstable to changes in the parameters of the model, which are roughly 20%, reduces strongly the number of predictions that are way-off. The method is then tested for robustness with data from 2021. We found that, for this data, only 1-2% of the one-week predictions were off by more than 50%. This increased to 3% for two-week predictions, and only for three-week predictions it reached 10%.

LTBI: latent tuberculosis infection or lasting immune responses to M. tuberculosis? A TBNET consensus statement.

Tuberculosis control relies on the identification and preventive treatment of individuals who are latently infected with Mycobacterium tuberculosis. However, direct identification of latent tuberculosis infection is not possible. The diagnostic tests used to identify individuals latently infected with M. tuberculosis, the in vivo tuberculin skin test and the ex vivo interferon-gamma release assays (IGRAs), are designed to identify an adaptive immune response against, but not necessarily a latent infection with, M. tuberculosis. The proportion of individuals who truly remain infected with M. tuberculosis after tuberculin skin test or IGRA conversion is unknown. It is also uncertain how long adaptive immune responses towards mycobacterial antigens persist in the absence of live mycobacteria. Clinical management and public healthcare policies for preventive chemotherapy against tuberculosis could be improved, if we were to gain a better understanding on M. tuberculosis latency and reactivation. This statement by the TBNET summarises knowledge and limitations of the currently available tests used in adults and children for the diagnosis of latent tuberculosis infection. In summary, the main issue regarding testing is to restrict it to those who are known to be at higher risk of developing tuberculosis and who are willing to accept preventive chemotherapy.

Effectiveness and safety of a treatment regimen based on isoniazid plus vaccination with Mycobacterium tuberculosis cells' fragments: field-study with naturally Mycobacterium caprae-infected goats.

The identification of a herd of goats with tuberculosis let us test a new treatment regimen against latent tuberculosis infection (LTBI). Using large animal experimental models allows a better approach to understanding human tuberculosis according to immunopathological parameters. Based on an initial study showing a correlation between the ESAT-6-specific interferon (IFN)-gamma secretion and the severity of pulmonary lesions, this parameter was used in combination with an X-ray examination to screen the animals to be included in the efficacy and safety studies. All the animals proved to be infected with Mycobacterium caprae. The efficacy study was run in animals distributed in three experimental groups according to treatment: untreated (CT), treated with isoniazid (INH), and treated with INH + RUTI (a vaccine based on M. tuberculosis cell fragments) inoculated twice. RUTI temporarily increased the IFN-gamma production after stimulating the peripheral blood with ESAT-6, purified protein derivative and RUTI in vitro. The INH chemotherapy reduced both pulmonary and extra pulmonary affectation, but not disease in pulmonary lymph nodes. The addition of RUTI may have decreased extrapulmonary disease further but had no benefit to lung or lung lymph-nodes itself. Safety studies showed that inoculation of RUTI caused a temporary increase of rectal temperature (1-2 degrees C) and local swelling, both adverse effects being well tolerated. Neither systemic toxicity nor mortality was induced by the vaccination. The control of goats' infection by the therapeutic regimen consisting in INH chemotherapy + RUTI as well as its safety, represented a further step towards testing its effects in human LTBI in a future.

A dynamic reinfection hypothesis of latent tuberculosis infection.

BACKGROUND: It has been traditionally postulated that individuals, once infected by Mycobacterium tuberculosis, will retain throughout their entire lifetime latent bacilli which will remain dormant in old lesions. This bacillus would then be the source of a later reactivation of active tuberculosis (TB), with the aid of resuscitation factors. Unfortunately, the presence of these bacilli can only be predicted by indirect immunological methods, such as the tuberculin skin test (TST) or T cell interferon-gamma release assays. Other evidence shows that a 9-month isoniazid treatment of TST+ individuals converting to TB reduces the incidence of TB by approximately 90%. QUESTIONS: Taking into account this widely accepted framework, I suggest that there are at least three relevant questions to answer: (1) How can dormant bacilli remain in the lungs for an entire lifetime, taking into account constant cellular turnover and the healing of damaged tissues? (2) What provides the resuscitation factor to dormant bacilli, assuming that these latent bacilli are indeed present inside old lesions? (3) Why can a 9-month treatment with isoniazid eliminate dormant bacilli? As isoniazid is active only against growing bacilli, and thus is only able to destroy them after reactivation of latent bacilli, this treatment should have to be provided for life if the traditionally accepted postulate is correct. HYPOTHESIS: For a better understanding of latent TB infection. I propose a hypothesis that describes a dynamic scenario of constant endogenous reinfection with M. tuberculosis which explains the efficacy of the current standard of treatment. If this hypothesis is true, new strategies for the management of TB may arise.

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.

Neutral-red reaction is related to virulence and cell wall methyl-branched lipids in Mycobacterium tuberculosis.

Searching for virulence marking tests for Mycobacterium tuberculosis, Dubos and Middlebrook reported in 1948 that in an alkaline aqueous solution of neutral-red, the cells of the virulent H37Rv M. tuberculosis strain fixed the dye and became red in color, whereas the cells of the avirulent H37Ra M. tuberculosis strain remained unstained. In the 1950 and 1960s, fresh isolates of M. tuberculosis were tested for this neutral-red cytochemical reaction and it was reported that they were neutral-red positive, whereas other mycobacteria of diverse environmental origins that were non-pathogenic for guinea pigs were neutral-red negative. However, neutral-red has not really been proven to be a virulence marker. To test if virulence is in fact correlated to neutral-red, we studied a clinical isolate of M. tuberculosis that was originally neutral-red positive but, after more than 1 year passing through culture mediums, turned neutral-red negative. We found that, in comparison to the original neutral-red positive strain, this neutral-red negative variant was attenuated in two murine models of experimental tuberculosis. Lipid analysis showed that this neutral-red negative natural mutant lost the capacity to synthesize pthiocerol dimycocerosates, a cell wall methyl-branched lipid that has been related to virulence in M. tuberculosis. We also studied the neutral-red of different gene-targeted M. tuberculosis mutants unable to produce pthiocerol dimycocerosates or other cell wall methyl-branched lipids such as sulfolipids, and polyacyltrehaloses. We found a negative neutral-red reaction in mutants that were deficient in more than one type of methyl-branched lipids. We conclude that neutral-red is indeed a marker of virulence and it indicates important perturbations in the external surface of M. tuberculosis cells.

[Origin and development of RUTI, a new therapeutic vaccine against Mycobacterium tuberculosis infection]. / Origen y desarrollo de RUTI, una nueva vacuna terapéutica contra la infección por Mycobacterium tuberculosis.

This article reviews the pathophysiology of the latent form of Mycobacterium tuberculosis along with its natural history and progression in infected tissues. The proposed hypotheses regarding the relationship between M tuberculosis and the associated immune response, the cause of granuloma necrosis, the tolerance of a certain concentration of the bacillus in host tissues, the constant turnover of cells in the lung, and the effect of chemotherapy form the basis for the design of the therapeutic vaccine RUTI against latent M tuberculosis infection. This vaccine is generated from detoxified M tuberculosis cell fragments that facilitate a balanced T helper (Th) 1/Th2/Th3 response to a wide range of antigens along with intense antibody production. Treatment with RUTI following chemotherapy has been demonstrated to be effective in experimental models in mice and guinea pigs and does not exhibit toxicity.

Rapid detection of Mycobacterium tuberculosis in respiratory specimens, blood and other non-respiratory specimens by amplification of rRNA.

SETTING: Diagnostic methods employing gene technology based on amplification of DNA or RNA are expected to improve the speed, sensitivity, and specificity of Mycobacterium tuberculosis detection. The Amplified Mycobacterium Tuberculosis Direct Test (AMTDT) enables the amplification and detection of M. tuberculosis rRNA directly from respiratory specimens. OBJECTIVE: To evaluate the performance of the AMTDT in direct detection of M. tuberculosis in respiratory specimens, blood and other clinical samples, and to compare this method with conventional culture and staining techniques. DESIGN: A total of 554 samples from 450 patients were examined in this study. All clinical specimens (with the exception of bone marrow aspirates and blood samples) were digested and decontaminated with sodium dodecyl (lauryl) sulfate (SDS)-NaOH. Bone marrow aspirates and blood samples were treated with 10% SDS. All processed samples were stained by auramine-rhodamine fluorochrome and inoculated onto Löwenstein-Jensen and Coletsos solid media, and into BACTEC-12B medium. In addition, the blood samples were inoculated into BACTEC 13A medium. The AMTDT was performed according to manufacturer's instructions. In those cases where discrepant results were obtained for AMTDT and cultures, patients' clinical data and other microbiological results were evaluated. RESULTS: The sensitivity, specificity, and positive and negative predictive values for AMTDT were 87.5, 100, 100, and 96.7%, respectively, in respiratory specimens and 86.8, 100, 100, and 92.8%, respectively, in non-respiratory specimens. The differences in sensitivity of these two groups of specimens were not highly statistically significant (P > 0.005). CONCLUSION: The sensitivity and specificity of the AMTDT were satisfactory for detection of M. tuberculosis in all types of clinical samples. Some minor changes in assay format and laboratory protocols may increase the sensitivity of the AMTDT without adversely affecting its specificity.

Double-blind, randomized, placebo-controlled Phase I Clinical Trial of the therapeutical antituberculous vaccine RUTI.

A Phase I interventional Clinical Trial was performed with a potential tuberculosis vaccine, based on detoxified cellular fragments of M. tuberculosis, named RUTI. The objective was to evaluate the safety profile and T-cell immune responses over a 6-month period following subcutaneous inoculation. The double-blind, randomized and placebo-controlled trial was conducted in healthy volunteers, all recruited at one site. RUTI, at each of the four tested doses, starting from 5microg and going up to 200microg, and placebo were inoculated to groups of 4 and 2 volunteers respectively, consecutively. RUTI appeared to be well tolerated as judged by local and systemic clinical evaluation, though vaccine dose dependent local adverse reactions were recorded. T-cell responses of blood lymphocytes to PPD and a number of antigen subunits were elevated, when compared with controls subjects. These results support the feasibility of future evaluation, to be targeted at subjects with latent tuberculosis infection (LTBI).

Extended safety studies of the attenuated live tuberculosis vaccine SO2 based on phoP mutant.

Safety is one of the main concerns for attenuated live vaccine candidates. Here we extend the stability and attenuation studies of the promising tuberculosis vaccine candidate based on Mycobacterium tuberculosis phoP mutant strain, SO2. Stability of the phoP mutation was tested after sub-culturing SO2 strain for 6 months in laboratory media and also after 3 months of infection in SCID mice. Results showed no reversion of the phoP mutation either in vitro or in vivo. In addition, SO2 was fully sensitive to four major first-line antituberculous drugs against tuberculosis. Safety and toxicity studies were performed in guinea pigs. Animals were infected with a quantity of SO2 equivalent to 50 vaccination doses (2.5x10(6) CFUs) and weight was monitored for 6 months. All animals survived and no histological lesions were found, showing full attenuation of SO2. Studies in a post-exposure model of guinea pigs and mice, previously infected with M. tuberculosis, were performed and no toxicity effects were found after inoculation of SO2. All these results together confirm that SO2 has a secure safety profile that encourages its use in clinical trials.

Usefulness of acr expression for monitoring latent Mycobacterium tuberculosis bacilli in 'in vitro' and 'in vivo' experimental models.

Real-time RT-PCR was used to quantify the expression of genes possibly involved in Mycobacterium tuberculosis latency in in vitro and murine models. Exponential and stationary phase (EP and SP) bacilli were exposed to decreasing pH levels (from 6.5 to 4.5) in an unstirred culture, and mRNA levels for 16S rRNA, sigma factors sigA,B,E,F,G,H and M, Rv0834c, icl, nirA, narG, fpbB, acr, rpoA, recA and cysH were quantified. The expression of acr was the one that best correlated with the CFU decrease observed in SP bacilli. In the murine model, the expressions of icl, acr and sigF tended to decrease when bacillary counts increased and vice versa. Values from immunodepressed mice (e.g. alpha/beta T cells, TNF, IFN-gamma and iNOs knock out strains), with accelerated bacillary growth rate, confirmed this fact. Finally, the expression of acr was maintained in mice following long-term treatment with antibiotics. The quantification of acr expression could be useful for monitoring the presence of latent bacilli in some murine models of tuberculosis.

On the nature of Mycobacterium tuberculosis-latent bacilli.

Mycobacterium tuberculosis-latent bacilli are microorganisms that adapt to stressful conditions generated by the infected host against them. By slowing metabolism or becoming dormant, they may counterbalance these conditions and appear as silent to the immune system. Moreover, the dynamic turnover of the infected cells provokes a constant reactivation of the latent bacilli when the environmental conditions are favourable, or an activation after being dormant in necrotic and fibrotic lesions for a long period of time. Since there is no in vivo nor in vitro evidence for quick resuscitation of dormant bacilli, the current authors strongly favour the possibility that latent tuberculosis infection can be maintained for no longer than approximately 10 yrs, which is, nowadays, a time period very close to that considered for "primary" tuberculosis. This concept may also be helpful for newer epidemiological considerations regarding the real impact of reinfection in tuberculosis.

Evolution of granulomas in lungs of mice infected aerogenically with Mycobacterium tuberculosis.

Aerogenous infection of C57Bl/6 mice with a virulent strain of Mycobacterium tuberculosis (CL 511) leads to the formation of primary granulomas in the lung where neutrophils, macrophages and subsequently, lymphocytes accumulate progressively around an initial cluster of infected macrophages. The spread of infection through the lung parenchyma gives rise to secondary granulomas featuring numerous lymphocytes that surround a small number of infected macrophages. Afterwards, foamy macrophages add an outer layer to the granulomas, which characteristically respect the pulmonary interstitium and remain confined within the alveolar spaces. This feature, in conjunction with the constant presence of M. tuberculosis in the products of broncho-alveolar lavage, suggests that the upward bronchial migration of infected macrophages may contribute significantly to pulmonary dissemination of mycobacterial infection. The latter would be in agreement with the persistence of chronic pulmonary infection in spite of a concomitant strong T helper 1 cell response.

Comparative evaluation of initial and new versions of the Gen-Probe Amplified Mycobacterium Tuberculosis Direct Test for direct detection of Mycobacterium tuberculosis in respiratory and nonrespiratory specimens.

We evaluated the initial version of the Amplified Mycobacterium Tuberculosis Direct Test (Gen-Probe) (AMTDT 1) and the new version of AMTDT (AMTDT 2) for the detection of Mycobacterium tuberculosis directly from respiratory and nonrespiratory samples and compared the results with those of culture and staining methods. The assays were applied to 410 respiratory and 272 nonrespiratory samples collected from 515 patients. The combination of the culture results and clinical diagnosis was considered to be the "gold standard." Ninety-five respiratory specimens were collected from 67 patients with a diagnosis of pulmonary tuberculosis (TB) and 68 nonrespiratory specimens were collected from 61 patients with a diagnosis of extrapulmonary TB. With respiratory specimens, the sensitivity, specificity, and positive and negative predictive values were 83, 100, 100, and 96%, respectively, for AMTDT 1 and 94.7, 100, 100, and 98.4%, respectively, for AMTDT 2. With nonrespiratory specimens, the sensitivity, specificity, and positive and negative predictive values were 83, 100, 100, and 94%, respectively, for AMTDT 1 and 86.8, 100, 100, and 98.4%, respectively, for AMTDT 2. The overall results of AMTDT 1 and AMTDT 2 were concordant for 97% (661 of 682) of the samples. Statistically significant differences in sensitivities were found between AMTDT 1 and AMTDT 2 with respiratory specimens. It was concluded that although both nucleic acid amplification methods are rapid, sensitive, and specific for the detection of M. tuberculosis complex in all types of clinical samples, AMTDT 2 appeared to be more sensitive than AMTDT 1 when applied to smear-negative specimens. In contrast AMTDT 2 is more susceptible than AMTDT 1 to inhibitory substances in the amplification reaction. The turnaround time of AMTDT 2 is shorter (3.5 h) than that for AMTDT 1 (5 h).

The intravenous model of murine tuberculosis is less pathogenic than the aerogenic model owing to a more rapid induction of systemic immunity.

The detection of mRNA in the murine model of tuberculosis for key cytokines involved in protective immunity in the lung tissues revealed a much faster emergence of the interferon (IFN)-gamma response in the intravenous route than in the aerosol route of inoculation. This slower response in the lungs was associated with a stronger inflammatory response, resulting in large granulomatous structures and eventual tissue damage.

Risk factors for lower airway bacterial colonization in chronic bronchitis.

The aim of this study was to determine the prevalence and risk factors for lower airway bacterial colonization (LABC) in stable chronic bronchitis (CB). Forty-one outpatients with CB were enrolled in the study (age 63.8+/-9.1 yrs (mean+/-SD); forced expiratory volume in one second (FEV1)/forced vital capacity (FVC) 62.8+/-11.2; current/former smokers 24/17). All patients had normal chest radiographs and an indication for performing fibreoptic bronchoscopy (pulmonary nodule, remote haemoptysis). The protected specimen brush (PSB) was used for bacterial sampling, and concentrations > or = 1,000 colony-forming units (cfu) x mL(-1) were considered positive for LABC. The repeatability of the procedure in CB was assessed in a random subsample of 18 subjects. A 72.2% quantitative agreement was found in the repeatability assessment of the PSB technique. Positive PSB cultures, obtained in 9 out of 41 (22%) patients, mainly yielded Haemophilus influenzae. The logistic regression model, used to determine which variables were related to colonization, showed that LABC was associated with current smoking (odds ratio (OR) 9.83, confidence interval (CI) 1.16-83.20) and low FVC (OR 0.73, CI 0.65-0.81). Age and FEV1 were not related to LABC. It was concluded that the prevalence of LABC in stable CB is high (22%), and current smoking is an important risk factor.

[Invasive Streptococcus agalactiae infections at a general university hospital over a 10-year period]. / Infecciones invasivas por Streptococcus agalactiae en un hospital general universitario durante un período de 10 años.

BACKGROUND: Streptococcus agalactiae is a known causal agent of neonatal meningitis, sepsis and puerperal infections. The incidence of invasive infections caused by Streptococcus agalactiae has increased in recent years in non gestating adults: in the elderly, patients receiving prolonged steroid treatment or those with chronic immunosuppressive diseases. The clinical and epidemiological characteristics and risk factors associated to invasive infections caused by S. agalactiae were analyzed. METHOD: A retrospective study was undertaken in patients with invasive disease by S. agalactiae attended in the Hospital Universitari Germans Trias i Pujol in Badalona (Barcelona), Spain, from 1983 to 1993. RESULTS: S. agalactiae was isolated in 51 patients including 13 (25%) neonates. Three patients presented invasive puerperal infection. Thirty-five adult patients with a mean age of 62 years presented invasive disease. Infection involved bacteremia in 26 (74.2%) patients. S. agalactiae was isolated in the ascitic fluid of 4 patients with liver cirrhosis with spontaneous bacterial peritonitis (one with bacteriemia) and in the peritoneal exudate of two patients with peritonitis secondary to intestinal perforation. Of 5 patients with septic arthritis, 3 involved bacteremia. Two patients presented empyema by S. agalactiae. Mortality was 28%, being directly related with infection in 4 cases (7.8%). CONCLUSIONS: Without taking pregnant women into account, 68% of the cases of invasive infections by S. agalactiae were observed in adults with associated base disease, with liver cirrhosis, neoplasms and diabetes mellitus being the most frequent. Advanced age was also found to be an important predisposing factor.

Towards a 'human-like' model of tuberculosis: intranasal inoculation of LPS induces intragranulomatous lung necrosis in mice infected aerogenically with Mycobacterium tuberculosis.

It is well known that one of the differences between murine and human tuberculosis is the lack of intragranulomatous necrosis in the former. The aim of this study was to create a feasible and reproducible model of an experimental model of murine tuberculosis in which this necrosis should be present. Considering the Shwartzman reaction as a possible explanation for intragranulomatous necrosis in human tuberculosis, C57Bl/6 mice, infected aerogenically with a virulent strain of Mycobacterium tuberculosis, were intranasally inoculated with lipopolysaccharide (LPS) on day 19 postinfection (p.i.). Twenty-four hours later, neutrophils infiltrated the lung parenchyma in a significant level, and 10 days after necrosis could be detected in the centres of primary granulomas, that showed scanty macrophages and large amounts of collagen on an eosinophilic background. On the other hand, a significant decrease in the concentration of colony forming units (CFU) could be appreciated 24 h after the LPS inoculation. Afterwards, nonbronchogenic spreading of granulomas increased and higher levels of interferon (IFN)-gamma mRNA were detected. These results lend support to the Shwartzman reaction as the origin of the intragranulomatous necrosis in the M. tuberculosis infection, and provides a useful tool to improve experimental murine models in tuberculosis.