An update on Mycobacterium tuberculosis lipoproteins.

Almost 3% of the proteins of Mycobacterium tuberculosis (M. tuberculosis), the main causative agent of human tuberculosis, are lipoproteins. These lipoproteins are characteristic of the mycobacterial cell envelope and participate in many mechanisms involved in the pathogenesis of M. tuberculosis. In this review, the authors provide an updated analysis of M. tuberculosis lipoproteins and categorize them according to their demonstrated or predicted functions, including transport of compounds to and from the cytoplasm, biosynthesis of the mycobacterial cell envelope, defense and resistance mechanisms, enzymatic activities and signaling pathways. In addition, this updated analysis revealed that at least 40% of M. tuberculosis lipoproteins are glycosylated.

International Analgesia and Sedation Weaning and Withdrawal Practices in Critically Ill Adults: The Adult Iatrogenic Withdrawal Study in the ICU.

OBJECTIVES: Iatrogenic withdrawal syndrome (IWS) associated with opioid and sedative use for medical purposes has a reported high prevalence and associated morbidity. This study aimed to determine the prevalence, utilization, and characteristics of opioid and sedative weaning and IWS policies/protocols in the adult ICU population. DESIGN: International, multicenter, observational, point prevalence study. SETTING: Adult ICUs. PATIENTS: All patients aged 18 years and older in the ICU on the date of data collection who received parenteral opioids or sedatives in the previous 24 hours. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: ICUs selected 1 day for data collection between June 1 and September 30, 2021. Patient demographic data, opioid and sedative medication use, and weaning and IWS assessment data were collected for the previous 24 hours. The primary outcome was the proportion of patients weaned from opioids and sedatives using an institutional policy/protocol on the data collection day. There were 2,402 patients in 229 ICUs from 11 countries screened for opioid and sedative use; 1,506 (63%) patients received parenteral opioids, and/or sedatives in the previous 24 hours. There were 90 (39%) ICUs with a weaning policy/protocol which was used in 176 (12%) patients, and 23 (10%) ICUs with an IWS policy/protocol which was used in 9 (0.6%) patients. The weaning policy/protocol for 47 (52%) ICUs did not define when to initiate weaning, and the policy/protocol for 24 (27%) ICUs did not specify the degree of weaning. A weaning policy/protocol was used in 34% (176/521) and IWS policy/protocol in 9% (9/97) of patients admitted to an ICU with such a policy/protocol. Among 485 patients eligible for weaning policy/protocol utilization based on duration of opioid/sedative use initiation criterion within individual ICU policies/protocols 176 (36%) had it used, and among 54 patients on opioids and/or sedatives ≥ 72 hours, 9 (17%) had an IWS policy/protocol used by the data collection day. CONCLUSIONS: This international observational study found that a small proportion of ICUs use policies/protocols for opioid and sedative weaning or IWS, and even when these policies/protocols are in place, they are implemented in a small percentage of patients.

Assessment of Tuberculosis Biomarkers in Paratuberculosis-infected Cattle.

Introduction: Mycobacterium bovis and Mycobacterium avium subsp. paratuberculosis, respectively the causative agents of bovine tuberculosis (bTB) and bovine paratuberculosis (PTB), share a high number of antigenic proteins. This characteristics makes the differential diagnosis of the diseases difficult. The interferon gamma (IFN-γ), C-X-C motif chemokine ligand 10 (CXCL10), matrix metallopeptidase 9 (MMP9), interleukin 22 (IL-22) and thrombospondin 1 (THBS1) bovine genes have already been shown to be accurate transcriptional biomarkers of bTB. In order to improve the diagnosis of bTB and PTB, in the present study we evaluated the risk of false positivity of these bTB biomarkers in cattle with PTB. Material and Methods: The transcription of these genes was studied in 13 PTB-infected cattle, using Mycobacterium avium subsp. paratuberculosis (MAP)-stimulated peripheral blood mononuclear cells (PBMC). Results: Overall, the levels of IFN-γ, CXCL10, MMP9 and IL-22 transcripts in MAP-stimulated PBMC failed to differentiate animals with PTB from healthy animals. However, as bTB-afflicted cattle do, the MAP-infected group also displayed a lower level of THBS1 transcription than the non-infected animals. Conclusion: The results of this study add new specificity attributes to the levels of transcription of IFN-γ, CXCL10, MMP9 and IL-22 as biomarkers for bTB.

Replication and transmission features of two experimental vaccine candidates against bovine tuberculosis subcutaneously administrated in a murine model.

Cattle vaccination is an attractive approach in compliance with control and eradication programs against Bovine Tuberculosis (bTB). Today, there is no anti bTB vaccine licensed. Two vaccine candidates, MbΔmce2 and MbΔmce2-phoP previously designed were evaluated in BALB/c mice, including the parental M. bovis NCTC10772 and a M. bovis hypervirulent Mb04-303 strains as controls. Sentinel mice (non-inoculated) cohoused with subcutaneous inoculated mice. Persistence, visible tuberculosis lesions (VTL) in lungs and spleens and bacillary load were investigated subcutaneously delivered at 60 and 90 days after inoculation (dpi) as well as their potential transmission to naïve mice. While a 100% survival was observed at 90 dpi without VTL in all groups, transmission was not evidenced in the sentinels mice. Vaccine candidates and control strains were isolated from the spleen of all inoculated mice, while Mb04-303 was isolated from the lungs of one inoculated mouse. Vaccine candidate's attenuation considering survival, lung bacillary load and VTL was confirmed, administrated by the subcutaneous route. Future experiments are necessary to demonstrate whether the persistence of both mutants in the spleen, with low CFU, remains over time to increase the potential increasing risk of dissemination to organs and subsequent transmission to other animals by airborne or other routes.

H65 fusion protein fails to improve the protection of a rationally attenuated live vaccine candidate against bovine tuberculosis in a mouse model of tuberculosis.

Background: The fusion protein H65, composed of Mycobacterium tuberculosis (TB) ESX-secreted antigens, has improved the bacillus Calmette-Guerin-induced immune protection in a mouse model of bovine TB when formulated in the liposomal adjuvant CAF01. In this study, we aimed to evaluate the protective efficacy of an attenuated Mycobacterium bovis strain - a mutant in mce2 and phoP genes - combined with H65+CAF01 immunization. We evaluated the protection of MbΔmce2-phoP alone or combined with H65+CAF01 against M. bovis challenge in mice. Methods: Groups of BALBc mice were inoculated with the vaccine candidates or phosphate buffered saline (PBS), and 6 weeks after the last immunization, the animals were aerogenically challenged with virulent M. bovis. Bacterial load in organs was counted after 45 days of the challenge. One-way analysis of variance and Bonferroni's posttest were used for statistical analysis. Results: All vaccinated mice showed reduced bacterial loads in lungs compared to unvaccinated animals. However, the protection level was similar between vaccinated groups. Conclusions: The MbΔmce2-phoP strain combined with three doses of H65+CAF01 induced equivalent protection than the MbΔmce2-phoP strain alone. Thus, the use of combined vaccination strategies requires a careful analysis of the potential interactions of each of their components with the host's immune system.

The subunit vaccine H65 + CAF01 increased the BCG- protection against Mycobacterium bovis infection in a mouse model of bovine tuberculosis.

H65, a fusion protein of three pairs of ESX-secreted antigens of Mycobacterium tuberculosis and Mycobacterium bovis, formulated with the liposomal adjuvant CAF01 has been shown to confer protection against M. tuberculosis infection in mice. In this study, we evaluated the impact of combining BCG with H65 + CAF01 immunization in a M. bovis mouse model of infection. We found that a BCG-H65 + CAF01/ H65 + CAF01 prime-boost scheme induced higher protection than BCG and H65 + CAF01 alone. Altogether, H65 antigen formulated in liposomal adjuvant improved the BCG-induced immune protection, thus making this vaccine strategy a promising tool to control bovine tuberculosis.

Elimination of ESAT-6 and CFP-10 from a candidate vaccine against bovine tuberculosis impaired its protection efficacy in the BALBc mouse model.

Background: Bovine tuberculosis (bTB) is a zoonotic disease caused by Mycobacterium bovis that mainly affects cattle. Although vaccination is the most effective strategy to control bTB, it may interfere with the diagnosis of the infection. Therefore, ancillary tests to differentiate vaccinated from infected animals (DIVA) are essential in a cattle vaccination scenario. ESAT-6 and CFP-10 are the most promissory DIVA antigens. Method: In this study, we deleted esat6 and cfp10 genes from the M. bovis Δ mce2 live-attenuated vaccine candidate and evaluated its protection level against bTB in BALBc mice. Results: We found that the M. bovis strain mutant in mce2, esat-6 and cfp-10 failed to confer protection against virulent M. bovis challenge in a mouse model of tuberculosis. Conclusions: This result highlights the relevant role of ESAT-6 and CFP-10 in the induction of protective immune response against M. bovis infection and reveals the need of evaluating different strategies to compensate for the lack of these DIVA antigens in new vaccine formulations.

FasR Regulates Fatty Acid Biosynthesis and Is Essential for Virulence of Mycobacterium tuberculosis.

Mycobacterium tuberculosis, the etiologic agent of human tuberculosis, is the world's leading cause of death from an infectious disease. One of the main features of this pathogen is the complex and dynamic lipid composition of the cell envelope, which adapts to the variable host environment and defines the fate of infection by actively interacting with and modulating immune responses. However, while much has been learned about the enzymes of the numerous lipid pathways, little knowledge is available regarding the proteins and metabolic signals regulating lipid metabolism during M. tuberculosis infection. In this work, we constructed and characterized a FasR-deficient mutant in M. tuberculosis and demonstrated that FasR positively regulates fas and acpS expression. Lipidomic analysis of the wild type and mutant strains revealed complete rearrangement of most lipid components of the cell envelope, with phospholipids, mycolic acids, sulfolipids, and phthiocerol dimycocerosates relative abundance severely altered. As a consequence, replication of the mutant strain was impaired in macrophages leading to reduced virulence in a mouse model of infection. Moreover, we show that the fasR mutant resides in acidified cellular compartments, suggesting that the lipid perturbation caused by the mutation prevented M. tuberculosis inhibition of phagolysosome maturation. This study identified FasR as a novel factor involved in regulation of mycobacterial virulence and provides evidence for the essential role that modulation of lipid homeostasis plays in the outcome of M. tuberculosis infection.

DETERMINATION OF THE RESPONSE TO THE ATMOSPHERIC COSMIC RADIATION OF A NEUTRON DOSIMETER ASSISTED BY MONTE CARLO SIMULATION.

A TLD-based dosimeter of polyethylene-lead-polyethylene, was developed and characterized with Monte Carlo simulations, using the MCNPX code. This passive system for the determination of the ambient dose equivalent (H*(10)) for neutrons over a wide energy range can be used for the dosimetry of neutrons from atmospheric cosmic radiation, on the ground, and onboard aircraft. A method assisted by Monte Carlo simulations that improves the calibration of fast neutron dosimeters based on moderation and thermalization of the incident fast flux and the measurement of the thermal flux by a sensor, which respond mainly to thermal neutrons, is presented in this work. The H*(10) energy response of this dosimeter was obtained from simulations for monoenergetic neutrons from 10-10 to 104 MeV. The validation of the modeling was done with irradiations for ISO standard neutron fields of 241Am-Be, 252Cf and 252Cf(D2O) at Instituto de Radioproteção e Dosimetria (IRD, Brazil) and at CERN-EU high-energy reference field (CERF).

Gene Pool Subdivision of East African Sweetpotato Parental Material.

Sweetpotato [Ipomoea batatas (L.) Lam] breeding is important for food security and health in East Africa (EA), and a breeding platform in Uganda provides national researchers and breeders in EA with true seed. Our objectives were to characterize genetic relationships among parental material used at the EA breeding platform. There were 135 parents and six check clones analyzed using 31 simple sequence repeat primers. An average of 7.13 alleles per primer was found, and Jaccard similarity coefficients were in the range of 0.298 to 1.00 with a mean of 0.542. Unweighted pair group cluster analysis placed most African parents in two main subclusters showing no association with morphology or geographical origin. The subclusters were also supported by principal coordinate analysis, derivative analysis of principal components, and population structure simulations. The analyzed breeding material from EA was highly genetically variable, grouped in two distinct genetic pools, and suitable to study heterosis exploiting breeding schemes.

INVESTIGATION OF THE INFLUENCE OF THE POSITION INSIDE A SMALL AIRCRAFT ON THE COSMIC-RADIATION-INDUCED DOSE.

This article report the measurements on-board a small aircraft at the same altitude and around the same geographic coordinates. The measurements of Ambient Dose Equivalent Rate (H*(10)) were performed in several positions inside the aircraft, close and far from the pilot location and the discrimination between neutron and non-neutron components. The results show that the neutrons are attenuated close to fuel depots and the non-neutron component appears to have the opposite behavior inside the aircraft. These experimental results are also confronted with results from Monte Carlo simulation, obtained with the MCNPX code, using a simplified model of the Learjet-type aircraft and a modeling of the standard atmosphere, which reproduces the real energy and angular distribution of the particles. The Monte Carlo simulation agreed with the experimental measurements and shows that the total H*(10) presents small variation (around 1%) between the positions inside aircraft, although the neutron spectra present significant variations.

Evaluation of Mycobacterium bovis double knockout mce2-phoP as candidate vaccine against bovine tuberculosis.

In this study, a Mycobacterium bovis knockout strain in phoP-phoR and mce2 operons was tested as an antituberculosis experimental vaccine in animal models. The double mutant strain was significantly more attenuated than the wild type strain in inmunocompetent and inmunodeficient mice. Vaccination with the double mutant protected mice against challenge with a virulent M. bovis strain.

Assessment of Mycobacterium bovis deleted in p27-p55 virulence operon as candidate vaccine against tuberculosis in animal models.

A Mycobacterium bovis knockout in p27-p55 operon was tested as an antituberculosis experimental vaccine in animal models. The mutant MbΔp27-p55 was significantly more attenuated in nude mice than its parental strain but more virulent than BCG Pasteur. Challenge experiments in mice and guinea pigs using M. bovis or M. tuberculosis strains showed similar protection conferred by MbΔp27-p55 mutant than BCG in terms of pathology and bacterial loads in spleen but lower protection than BCG in lungs. When tested in cattle, MbΔp27-p55 did not induce IL-2 expression and induced a very low production of IFNγ, suggesting that the lack of P27/P55 reduces the capacity of M. bovis of triggering an adequate Th1 response.

The ochodaeidae of Argentina (coleoptera, scarabaeoidea).

The Ochodaeidae (Coleoptera: Scarabaeoidea) of Argentina are revised. Previously, two species of Ochodaeinae were known from the country, both in the genus Parochodaeus Nikolajev: Parochodaeus campsognathus (Arrow) and Parochodaeus cornutus (Ohaus). An additional 7 species of Parochodaeus from Argentina are described here as new. In addition, Gauchodaeus patagonicus, new genus and new species in the subfamilyChaetocanthinae, is described. This is the first record of the subfamily Chaetocanthinae in South America. Redescriptions, diagnoses, and maps are provided for each species. We also provide a key to genera and a key to species of Parochodaeus of Argentina. With this work, the number of ochodaeid species known from Argentina is increased from 2 to 10.

Impact of the deletion of the six mce operons in Mycobacterium smegmatis.

The Mycobacterium smegmatis genome contains six operons designated mce (mammalian cell entry). These operons, which encode membrane and exported proteins, are highly conserved in pathogenic and non-pathogenic mycobacteria. Although the function of the Mce protein family has not yet been established in Mycobacterium smegmatis, the requirement of the mce4 operon for cholesterol utilization and uptake by Mycobacterium tuberculosis has recently been demonstrated. In this study, we report the construction of an M. smegmatis knock-out mutant deficient in the expression of all six mce operons. The consequences of these mutations were studied by analyzing physiological parameters and phenotypic traits. Differences in colony morphology, biofilm formation and aggregation in liquid cultures were observed, indicating that mce operons of M. smegmatis are implicated in the maintenance of the surface properties of the cell. Importantly, the mutant strain showed reduced cholesterol uptake when compared to the parental strain. Further cholesterol uptake studies using single mce mutant strains showed that the mutation of operon mce4 was reponsible for the cholesterol uptake failure detected in the sextuple mce mutant. This finding demonstrates that mce4operon is involved in cholesterol transport in M. smegmatis.

Evaluation of pathogenesis caused in cattle and guinea pig by a Mycobacterium bovis strain isolated from wild boar.

BACKGROUND: In many regions of the world, wild mammals act as reservoir of Mycobacterium bovis, a situation that prevents the eradication of bovine tuberculosis. In order to observe whether a strain isolated from a wild boar, previously tested as highly virulent in a mice model, is also virulent in cattle, we performed cattle experimental inoculation with this strain RESULTS: Groups of Friesian calves were either infected with the wild boar strain M. bovis 04-303 or with the bovine strain NCTC10772 as a control. We found that antigen-specific IFN-γ release in whole blood samples occurred earlier in animals infected with M. bovis 04-303. Both M. bovis strains resulted in a positive skin test, with animals infected with the wild boar isolate showing a stronger response. These results and the presence of more severe organ lesions, with granuloma and pneumonic areas in cattle demonstrate that the wild boar isolate is more virulent than the NCTC10772 strain. Additionally, we tested the infectivity of the M. bovis strains in guinea pigs and found that M. bovis 04-303 had the highest pathogenicity. CONCLUSIONS: M. bovis strains isolated from wild boars may be pathogenic for cattle, producing TB lesions.

Role of P27 -P55 operon from Mycobacterium tuberculosis in the resistance to toxic compounds.

BACKGROUND: The P27-P55 (lprG-Rv1410c) operon is crucial for the survival of Mycobacterium tuberculosis, the causative agent of human tuberculosis, during infection in mice. P55 encodes an efflux pump that has been shown to provide Mycobacterium smegmatis and Mycobacterium bovis BCG with resistance to several drugs, while P27 encodes a mannosylated glycoprotein previously described as an antigen that modulates the immune response against mycobacteria. The objective of this study was to determine the individual contribution of the proteins encoded in the P27-P55 operon to the resistance to toxic compounds and to the cell wall integrity of M. tuberculosis. METHOD: In order to test the susceptibility of a mutant of M. tuberculosis H37Rv in the P27-P55 operon to malachite green, sodium dodecyl sulfate, ethidium bromide, and first-line antituberculosis drugs, this strain together with the wild type strain and a set of complemented strains were cultivated in the presence and in the absence of these drugs. In addition, the malachite green decolorization rate of each strain was obtained from decolorization curves of malachite green in PBS containing bacterial suspensions. RESULTS: The mutant strain decolorized malachite green faster than the wild type strain and was hypersensitive to both malachite green and ethidium bromide, and more susceptible to the first-line antituberculosis drugs: isoniazid and ethambutol. The pump inhibitor reserpine reversed M. tuberculosis resistance to ethidium bromide. These results suggest that P27-P55 functions through an efflux-pump like mechanism. In addition, deletion of the P27-P55 operon made M. tuberculosis susceptible to sodium dodecyl sulfate, suggesting that the lack of both proteins causes alterations in the cell wall permeability of the bacterium. Importantly, both P27 and P55 are required to restore the wild type phenotypes in the mutant. CONCLUSIONS: The results clearly indicate that P27 and P55 are functionally connected in processes that involve the preservation of the cell wall and the transport of toxic compounds away from the cells.

The South American dung beetle genus Ennearabdus Lansberge (Coleoptera: Scarabaeidae: Scarabaeinae: Eucraniini).

The South American endemic dung beetle genus Ennearabdus Lansberge is revised. Description, diagnosis and illustrations are presented for the only known species of the genus, E. lobocephalus (Harold). A lectotype is designated for Onthophagus lobocephalus Harold, the type species of Ennearabdus. The biology, biogeography, conservation status, and distribution based on the predictive distribution model of E. lobocephalus are also discussed.

A revision of the Argentinean endemic genus Eucranium Brullé (Coleoptera: Scarabaeidae: Scarabaeinae) with description of one new species and new synonymies.

The South American genus Eucranium Brullé has been revised and now includes six species: E. arachnoides Brullé, E. belenae Ocampo new species, E. cyclosoma Burmeister, E. dentifrons Guérin-Méneville, E. planicolle Burmeister, and E. simplicifrons Fairmaire. Eucranium pulvinatum Burmeister is a new junior synonym of Eucranium arachnoides Brullé, and Eucranium lepidum Burmeister is a new junior synonym of E. dentifrons Guérin-Méneville. The following lectotypes and neotypes are designated: Eucranium pulvinatum Burmeister, lectotype; Eucranium planicolle Burmeister, lectotype; Psammotrupes dentifrons Guérin-Méneville, neotype; and Eucranium lepidum Burmeister, neotype. Description of the genus and new species, diagnosis and illustrations, and distribution maps are provided for all species. A key to the species of this genus is provided, and the biology and conservation status of the species are discussed.