Insights into the genomic features and lifestyle of B1 subcluster mycobacteriophages.

Bacteriophages infecting Mycobacterium smegmatis mc2155 are numerous and, hence, are classified into clusters based on nucleotide sequence similarity. Analyzing phages belonging to clusters/subclusters can help gain deeper insights into their biological features and potential therapeutic applications. In this study, for genomic characterization of B1 subcluster mycobacteriophages, a framework of online tools was developed, which enabled functional annotation of about 55% of the previously deemed hypothetical proteins in B1 phages. We also studied the phenotype, lysogeny status, and antimycobacterial activity of 10 B1 phages against biofilm and an antibiotic-resistant M. smegmatis strain (4XR1). All 10 phages belonged to the Siphoviridae family, appeared temperate based on their spontaneous release from the putative lysogens and showed antibiofilm activity. The highest inhibitory and disruptive effects on biofilm were 64% and 46%, respectively. This systematic characterization using a combination of genomic and experimental tools is a promising approach to furthering our understanding of viral dark matter.

Coordination of carbon partitioning and photosynthesis by a two-component signaling network in Synechococcus elongatus PCC 7942.

Photosynthetic organisms need to balance the rate of photosynthesis with the utilization of photosynthetic products by downstream reactions. While such "source/sink" pathways are well-interrogated in plants, analogous regulatory systems are unknown or poorly studied in single-celled algal and cyanobacterial species. Towards the identification of energy/sugar sensors in cyanobacteria, we utilized an engineered strain of Synechococcus elongatus PCC 7942 that allows experimental manipulation of carbon status. We conducted a screening of all two-component systems (TCS) and serine/threonine kinases (STKs) encoded in S. elongatus PCC 7942 by analyzing phenotypes consistent with sucrose-induced relaxation of sink inhibition. We narrowed the candidate sensor proteins by analyzing changes observed after sucrose feeding. We show that a clustered TCS network containing RpaA, CikB, ManS and NblS are involved in the regulation of genes related to photosynthesis, pigment synthesis, and Rubisco concentration in response to sucrose. Altogether, these results highlight a regulatory TCS group that may play under-appreciated functions in carbon partitioning and energy balancing in cyanobacteria.

Comparing the Effect of Phenylephrine Bolus and Phenylephrine Infusion for Maintaining Arterial Blood Pressure During Cesarean Delivery Under Spinal Anesthesia: A Randomized Prospective Study.

INTRODUCTION:  Maternal hypotension is a common complication of spinal anesthesia in cesarean section and requires immediate intervention. Phenylephrine is most commonly used as a vasopressor agent for the treatment of hypotension due to subarachnoid block. Our aim was to compare the bolus dose of 50 µg of phenylephrine with a fixed infusion at 50 µg.min-1 of phenylephrine for maintaining arterial blood pressure during cesarean delivery. MATERIALS AND METHOD:  This was a prospective, randomized comparative study. One hundred normotensive pregnant females aged 18-35 years, body mass index 18-29kg.m2, American Society of Anesthesiologists (ASA) physical status classification II scheduled to undergo cesarean section (elective/emergency) under spinal anesthesia were randomly divided into two groups of 50 each. Group PB received a bolus dose of phenylephrine 50 µg after they developed hypotension i.e., a decrease in systolic blood pressure (SBP) ≥ 20% from the baseline. Similarly, patients in Group PI were administered prophylactic infusion using a syringe pump of phenylephrine 50 µg.min-1, started just after the administration of subarachnoid block. The phenylephrine infusion was continued either till the delivery of the baby or when SBP rises >20% above the baseline. Parameters like blood pressure, heart rate, and peripheral oxygen saturation were recorded. After the delivery of the baby, the neonatal APGAR score was assessed at one minute and five minutes. RESULTS: Demographic data were comparable in terms of demographic profile, duration of surgery, and ASA physical status classification between the groups. The heart rate was higher in Group PB compared to Group PI throughout the monitoring period (P<0.001). The fall in mean blood pressure was more in Group PB compared to Group PI till 18 minutes of surgery and was statistically significant (P<0.05). After 18 minutes of surgery, mean blood pressure stabilized and was comparable between the groups. Other variables like APGAR score at one minute and five minutes were comparable between the groups. Bradycardia and hypertension were more common in Group PI whereas hypotension, nausea, and vomiting were more common in group PB. CONCLUSION: We concluded that during cesarean section under spinal anesthesia, phenylephrine infusion provides better hemodynamic stability and APGAR score during the perioperative period.

Efficacy of Ultrasound-Guided Erector Spinae Plane Block in Percutaneous Nephrolithotomy.

Background Percutaneous nephrolithotomy (PCNL) is presently the preferred method for managing renal calculi. Visceral pain from the kidney and ureter and somatic pain from the incision site are the primary causes of immediate postoperative pain following PCNL. Poor pain control is associated with unwanted consequences such as patient discomfort, delayed recovery, and prolonged hospital stay. Recently, the erector spinae plane (ESP) block has been used in many thoracic and abdominal surgeries for the control of postoperative pain. In this study, we aimed to assess the effectiveness of the ultrasound-guided ESP block following PCNL. Methodology This was a prospective, double-blind, randomized controlled study including 60 patients who were scheduled for elective PCNL under general anesthesia. Patients were randomly divided into two groups. Group E underwent an ultrasound-guided ESP block with 20 mL of the local anesthetic mixture at the T-9 level unilaterally on the side of surgery, and group C was a sham group in which 20 mL of normal saline was injected on the side of surgery. Changes in postoperative pain score were the primary outcome, and the duration of analgesia, the total analgesic requirement in 24 hours, and patient satisfaction were the secondary outcomes. Results The demographic data of both groups were comparable. The Visual Analog Scale score was considerably lower in group E than in group C at two, four, six, and eight hours postoperatively. In group E, the mean analgesic duration was substantially longer than that in group C (8.87 ± 2.45 hours vs. 5.67 ± 1.58 hours, respectively). The tramadol requirement was higher in group C (286.67 ± 62.88 mg) than in group E (133.33 ± 47.95 mg) during the 24-hour postoperative period. At 12 hours, patient satisfaction was considerably higher in group E than in group C (6.73 ± 0.45 vs. 5.87 ± 0.35, respectively). Conclusions The ultrasound-guided ESP block provided efficient postoperative pain relief, prolonged duration of analgesia, and reduced tramadol intake after PCNL surgery.

Inhaled Adjunct Therapy with Second-Line Drug Candidates for Dose Reduction in Chemotherapeutic Regimens for Multi-drug-Resistant Tuberculosis.

Chemotherapy of multi-drug-resistant tuberculosis (TB) requires prolonged administration of multiple drugs. We investigated whether pulmonary delivery of minute doses of drugs, along with reduced oral doses of the same agents, would affect preclinical efficacy. We prepared dry powder inhalation (DPI) formulations comprising sutezolid (SUT), the second-generation pretomanid analog TBA-354 (TBA), or a fluorinated derivative of TBA-354 (32,625) in a matrix of the biodegradable polymer poly(L-lactide). We established formulation characteristics, doses inhaled by healthy mice, and preclinical efficacy in a mouse model of TB. Oral doses of 100 mg/kg/day or DPI doses of 0.25-0.5 mg/kg/day of drugs SUT, TBA-354, or 32,625 administered over 28 days were sub-optimally effective in reducing lung and spleen burden of Mycobacterium tuberculosis (Mtb) in infected mice. The addition of 0.25-0.5 mg/kg/day of SUT, TBA-354, or 32,625 as DPI to oral doses of 50 mg/kg/day was non-inferior in clearing Mtb from the lungs of infected mice. We concluded that adjunct therapy with inhaled second-line agents has the potential to reduce the efficacious oral dose.

Comparative Study Between Conventional Landmark Versus Ultrasound-Guided Paravertebral Block in Patients Undergoing Laparoscopic Cholecystectomy: A Randomized Controlled Study.

INTRODUCTION: Thoracic paravertebral block (TPVB) has emerged as an effective and safe regional technique for providing postoperative analgesia. We aimed to compare the ease and efficacy of conventional landmark and ultrasound-guided (USG) paravertebral blocks for postoperative analgesia in patients undergoing laparoscopic cholecystectomy. METHODS: This was a randomized controlled study. Seventy-six patients of either sex, age 18-40 years, body mass index (BMI) 18-29 kg/m2, American Society of Anesthesiologists physical status classifications I and II posted for elective laparoscopic cholecystectomy under general anesthesia were randomly allocated into two groups of 38 each. Patients in group A were administered a paravertebral block using the anatomical landmark technique (ALT), and group B using an ultrasound-guided paravertebral block in the sitting position. In both groups, 20 ml of 0.5% bupivacaine injection was administered at the T7 vertebral level on the right side. The primary outcome was the first-pass success rate. Secondary outcomes were the number of passes and attempts, duration of analgesia, visual analog scale (VAS) score for pain during 24 h postoperatively and complications if any, were recorded. RESULTS: No patients were excluded in the study. Demographic characteristics were comparable in both groups. The number of passes was less in group B (1.45±0.5) compared to group A (2.42±0.95) and was reported to be statistically significant (p = 0.001). The number of attempts was less in group B (1.00±0) as compared to group A (1.29±0.46) and was statistically significant (p = 0.001). The duration of analgesia was longer in group B (530.00±326.33 minutes) compared to group A (345.60±252.95 minutes) and was observed to be statistically significant (p<0.05). The VAS score was significantly lower in group B (1.87±0.78, 2.24 ±0.82) compared to group A (2.42±0.72, 3.13±1.07) at the second and fourth hours, respectively (p = 0.001).  Conclusion: We concluded that paravertebral block using an ultrasound-guided technique is more efficacious than the conventional landmark technique for postoperative analgesia in patients undergoing laparoscopic cholecystectomy.

Cohesin regulates alternative splicing.

Cohesin, a trimeric complex that establishes sister chromatid cohesion, has additional roles in chromatin organization and transcription. We report that among those roles is the regulation of alternative splicing through direct interactions and in situ colocalization with splicing factors. Degradation of cohesin results in marked changes in splicing, independent of its effects on transcription. Introduction of a single cohesin point mutation in embryonic stem cells alters splicing patterns, demonstrating causality. In primary human acute myeloid leukemia, mutations in cohesin are highly correlated with distinct patterns of alternative splicing. Cohesin also directly interacts with BRD4, another splicing regulator, to generate a pattern of splicing that is distinct from either factor alone, documenting their functional interaction. These findings identify a role for cohesin in regulating alternative splicing in both normal and leukemic cells and provide insights into the role of cohesin mutations in human disease.

Resiliencies of soil phosphorus fractions after natural summer fire are governed by microbial activity and cation availability in a semi-arid Inceptisol.

The unintended impact of natural summer fire on soil is complicated and rather less studied than its above-ground impact. Recognising the impact of a fire on silvopastoral soils and their resilience can aid in improving the management of silvopastoral systems. We studied the immediate (after 1 week (W)) and short-term (after 3 months (M)) recovery of different soil biological and chemical properties after the natural fire, with specific emphasis on phosphorus (P) dynamics. Soil samples were collected from four different layers (0-15, 15-30, 30-45, and 45-60 cm) of Morus alba, Leucaena leucocephala, and Ficus infectoria based silvopastoral systems. In the 0-15 cm soil layer, soil organic carbon (SOC) declined by ∼37, 42, and 30% after the fire in Morus-, Leucaena-, and Ficus-based systems, respectively within 1W of fire. However, after 3M of fire, Morus and Leucaena regained ∼6 and 11.5% SOC as compared to their status after 1W in the 0-15 cm soil layer. After 1W of the fire, soil nitrogen (N), sulfur (S), and potassium availability declined significantly at 0-15 cm soil layer in all systems. Iron and manganese availability improved significantly after 1W of the fire. Saloid bound P and aluminium bound P declined significantly immediately after the fire, increasing availability in all systems. However, calcium bound P did not change significantly after the fire. Dehydrogenase and alkaline phosphatase activity declined significantly after the fire, however, phenol oxidase and peroxidase activity were unaltered. Resiliencies of these soil properties were significantly impacted by soil depth and time. Path analysis indicated microbial activity and cationic micronutrients majorly governed the resilience of soil P fractions and P availability. Pasture yield was not significantly improved after the fire, so natural summer fire must be prevented to avoid loss of SOC, N, and S.

Lung Resident Memory T Cells Activated by Oral Vaccination Afford Comprehensive Protection against Pneumonic Yersinia pestis Infection.

A growing body of evidence has shown that resident memory T (TRM) cells formed in tissue after mucosal infection or vaccination are crucial for counteracting reinfection by pathogens. However, whether lung TRM cells activated by oral immunization with Yptb1(pYA5199) play a protective role against pneumonic plague remains unclear. In this study, we demonstrated that lung CD4+ and CD8+ TRM cells significantly accumulated in the lungs of orally Yptb1(pYA5199)-vaccinated mice and dramatically expanded with elevated IL-17A, IFN-γ, and/or TNF-α production after pulmonary Yersinia pestis infection and afforded significant protection. Short-term or long-term treatment of immunized mice with FTY720 did not affect lung TRM cell formation and expansion or protection against pneumonic plague. Moreover, the intratracheal transfer of both lung CD4+ and CD8+ TRM cells conferred comprehensive protection against pneumonic plague in naive recipient mice. Lung TRM cell-mediated protection was dramatically abolished by the neutralization of both IFN-γ and IL-17A. Our findings reveal that lung TRM cells can be activated via oral Yptb1(pYA5199) vaccination, and that IL-17A and IFN-γ production play an essential role in adaptive immunity against pulmonary Y. pestis infection. This study highlights an important new target for developing an effective pneumonic plague vaccine.

A review on cultural competency in medical education.

Cultural competency is a wide notion with a variety of academic bases and differing perspectives on how it should be implemented. While it is widely acknowledged that cultural competency should be an element of general practise, there is a paucity of literature in this area. It has been commonly claimed that cultural competency is a fundamental prerequisite for working well with persons from different cultural backgrounds. Medical students must learn how to connect successfully with patients from all walks of life, regardless of culture, gender, or financial background. Hence, National Medical Council (NMC) has included cultural competence as a course subject in the curriculum of medical education. The opportunities and concept of Competency Based Medical Education, the inclusion of cultural competency in medical course by NMC, various models and practice skill of cultural competence in medical education are discussed in this paper. This study will be useful to researchers who are looking at cultural competency as a research variable that influences study result.

Effect of Dietary Supplementation of Probiotic Aspergillus niger on Performance and Cecal Microbiota in Hy-Line W-36 Laying Hens.

This study aimed to investigate the role of the probiotic Aspergillus niger on the production performance, egg quality, and cecal microbial load of Clostridium perfringens, Salmonella spp., and Escherichia coli in Hy-Line W-36 laying hens. A total of 72, 45-week-old Hy-Line W-36 laying hens were randomly allocated to one of the three dietary treatments with six replicates, and each replicate had four individually caged laying hens (n = 6 and 4 hens/replicate). The hens in each treatment group were fed a corn and soybean meal diet (Control), a diet supplemented with bacitracin methylene disalicylate (BMD) at a rate of 495 mg/kg of feed (Positive Control), or a diet supplemented with Aspergillus niger (Probioist®) at a rate of 220 mg/kg of feed (Probiotic). Supplementing probiotics in the laying hen diet significantly increased egg production at weeks 3 and 6 compared with the Positive Control. Haugh unit, a measure of egg quality, was significantly higher in laying hens fed the probiotic diet compared with the Control or Positive Control at week 10. Furthermore, the Probiotic group had numerically lower cecal microbial loads of pathogenic bacteria (Clostridium perfringens, Salmonella spp., and Escherichia coli) compared with the Control and Positive Control groups. The results suggest that Aspergillus niger could be used as a probiotic to improve laying hen performance and egg quality.

TinoTranscriptDB: A Database of Transcripts and Microsatellite Markers of Tinospora cordifolia, an Important Medicinal Plant.

Tinospora cordifolia, commonly known as "Giloe" in India, is a shrub belonging to the family Menispermaceae. It is an important medicinal plant known for its antipyretic, anti-inflammatory, antispasmodic, and antidiabetic properties and is used in the treatment of jaundice, gout, and rheumatism. Despite its economic importance, the limited information related to its genomic resources prohibits its judicious exploitation through molecular breeding or biotechnological approaches. In this study, we generated a meta-transcriptome assembly of 43,090 non-redundant transcripts by merging the RNASeq data obtained from Roche 454 GS-FLX, and Illumina platforms, and report the first transcriptome-based database for simple sequence repeats and transcription factors ("TinoTranscriptDB" (Tinospora cordifolia Transcriptome Database)). We annotated 26,716 (62%) of the total transcripts successfully from National Center for Biotechnology Information non-redundant protein (NCBI-NR), gene ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), Swiss-Prot, and Pfam databases. This database contains information of 2620 perfect simple sequence repeats (P-SSRs) with a relative abundance of 340.12 (loci/Mb), and relative density of 6309.29 (bp/Mb). Excluding mono-nucleotides, the most abundant SSR motifs were tri-nucleotides (54.31%), followed by di-nucleotides (37.51%), tetra-nucleotides (4.54%), penta-nucleotides (3.16%) and hexa-nucleotides (0.45%). Additionally, we also identified 4,311 transcription factors (TFs) and categorized them into 55 sub-families. This database is expected to fill the gap in genomic resource availability in T. cordifolia and thus accelerate molecular breeding and related functional and other applied studies aimed towards genetic improvements of T. cordifolia and related species.

Anesthetic Stability of Propofol, Dexmedetomidine, and Isoflurane by Measuring Bispectral Index (BIS) and Hemodynamic Indices: A Comparative Study.

Background Hemodynamic changes and anesthetic awareness occurring during surgery under general anesthesia is a great concern for both surgeon and anesthesiologist. Maintenance of the adequate depth of anesthesia throughout the intraoperative period is important in maintaining hemodynamic stability, preventing intraoperative awareness, and avoiding postoperative recall. Aim This study aims to predict the anesthetic stability of propofol, dexmedetomidine, and isoflurane by measuring bispectral index (BIS) and hemodynamic indices. Materials and methods This is a prospective comparative study. Sixty patients of either sex, aged 18-60 years, with American Society of Anesthesiologist (ASA) physical status classification I and II, undergoing elective surgical procedures requiring general anesthesia were allocated into three groups of 20 each. Patients in each group were administered standard general anesthesia with routine hemodynamic monitoring along with BIS, and values were recorded at baseline and thereafter at every five-minute interval for the duration of surgery. Anesthesia was maintained in Group P using a bolus dose of propofol 1 milligram.kg-1 for 10 minutes followed by propofol infusion 50-75 microgram.kg-1.minute-1, Group D with a bolus dose of dexmedetomidine 1 microgram.kg-1 for 10 minutes followed by infusion 0.2-0.7 microgram.kg-1.hour-1, and Group I with isoflurane at 1 minimum alveolar concentration (MAC) for 10 minutes and then maintained between 0.5 MAC and 1.5 MAC until the duration of surgery. To maintain the surgical plane of anesthesia, the BIS score was monitored between 40 and 65. The quantitative variables were expressed as mean±SD and compared between groups using Student's unpaired t-test. Data analysis was done using SPSS Statistics for Windows version 20.0 (IBM Corp., Armonk, NY, USA). A p-value of <0.05 was considered statistically significant. Results During intergroup comparison among study drugs, the mean BIS values were statistically significant among the groups (p<0.05). Hemodynamic indices were significantly better maintained in the dexmedetomidine group as compared to the isoflurane and propofol groups throughout the intraoperative period (p<0.05). Conclusion Dexmedetomidine is better than propofol and isoflurane in maintaining the BIS score and hemodynamic parameters during the intraoperative period.

Randomized Comparative Study Between Bilateral Erector Spinae Plane Block and Transversus Abdominis Plane Block Under Ultrasound Guidance for Postoperative Analgesia After Total Abdominal Hysterectomy.

INTRODUCTION: Ultrasound-guided erector spinae plane (ESP) block has emerged as an effective and safe analgesic regional technique and it also provides visceral pain relief. Our aim was to compare the analgesic efficacy of ESP block over transversus abdominis plane (TAP) block under ultrasound guidance following a total abdominal hysterectomy. METHODS: This was a prospective, randomized, comparative study. Thirty females posted for elective open total abdominal hysterectomy under general anesthesia were randomly allocated into two groups. Ultrasound-guided ESP block was applied in group E at the T-9 level bilaterally. The study solution was prepared by mixing 20 ml of 0.5% bupivacaine plus 10 ml of 2% lignocaine and 1 ml (50mcg) of fentanyl and 9 ml of normal saline forming total 40 ml of which 20 ml was injected on each side. Group T received ultrasound-guided TAP block with 20 ml of study solution bilaterally. The study solution was prepared similarly by mixing 20 ml of 0.5% bupivacaine plus 10 ml of 2% lignocaine and 1 ml (50mcg) of fentanyl and 9 ml of normal saline (total 40 ml) of which 20 ml was injected into each side. Tramadol 100mg iv was given as rescue analgesia whenever NRS ≥ 4 or on-demand in the postoperative period. The primary outcome was changes in a numerical rating scale (NRS) pain score postoperatively between two groups in 24 h, duration of analgesia and total rescue analgesic required during 24 h. The secondary outcome was patient satisfaction level and side effects if any. RESULTS: Demographic data were comparable in both groups. The NRS pain score was significantly lower ​​​​in group E than in group T at second, third, fourth, fifth (p < 0.001) and at sixth hour (p < 0.05) postoperatively. The mean duration of analgesia was significantly more in Group E (4.73±0.7 h) compared to group T (2.60±0.51 h) (p < 0.001). The tramadol consumption was seen significantly more in 24 h in group T (233.33±25.82 mg) than in group E (193.33±17.59 mg). Patient satisfaction score was significantly higher at 12 h with mean value of 6.07±0.26 in group E compared to 3.40±0.91 in group T. CONCLUSION: We conclude that ultrasound-guided ESP block provide better postoperative pain control and prolonged duration of analgesia with less tramadol consumption compared to ultrasound-guided TAP block in patients after total abdominal hysterectomy.

Modified Bain's Circuit as an Alternate to Non-invasive Ventilation in COVID 19.

Introduction COVID-19 is a pandemic that severely affects the lungs. Symptomatically affected individuals often become severely hypoxic, requiring non-invasive ventilation. The scarcity of resources in resource-compromised countries like India led to the adoption of novel strategies like using Bain's circuit for assisting spontaneous ventilation. This study compares the outcome when a standard circuit is replaced with a shortened Bain's circuit. Aims and objectives To compare shortened Bain's circuit and bilevel positive airway pressure (BiPAP) in spontaneously ventilated COVID 19 patients with regards to effects on hemodynamic stability and efficacy of ventilation using blood gas analysis. Methodology Twenty-four COVID patients aged between 35-70 years, requiring non-invasive ventilation but not tolerating BiPAP or not improving on BiPAP were enrolled in the study. Baseline heart rate and arterial blood gases (ABG) were recorded. Patients were then ventilated using shortened Bain's circuit. Heart rate and ABG were then recorded two hours after ventilation. Results Hemodynamic and blood gas parameters were comparable between the two groups at baseline and on BiPAP. Group A showed better hemodynamic and blood gas profiles compared to group B, but the difference was not statistically significant because of small sample size. Conclusion Shortened Bain's circuit may be a viable alternative to non-invasive ventilation in spontaneously breathing hypoxic patients with efficacy comparable to a standard Bain's circuit and reduced chances of carbon dioxide retention. Studies with a larger sample size are needed to further validate the conclusion.

Lethal synergy between SARS-CoV-2 and Streptococcus pneumoniae in hACE2 mice and protective efficacy of vaccination.

Secondary infections are frequent complications of viral respiratory infections, but the potential consequence of SARS-CoV-2 coinfection with common pulmonary pathogens is poorly understood. We report that coinfection of human ACE2-transgenic mice with sublethal doses of SARS-CoV-2 and Streptococcus pneumoniae results in synergistic lung inflammation and lethality. Mortality was observed regardless of whether SARS-CoV-2 challenge occurred before or after establishment of sublethal pneumococcal infection. Increased bacterial levels following coinfection were associated with alveolar macrophage depletion, and treatment with murine GM-CSF reduced numbers of lung bacteria and pathology and partially protected from death. However, therapeutic targeting of IFNs, an approach that is effective against influenza coinfections, failed to increase survival. Combined vaccination against both SARS-CoV-2 and pneumococci resulted in 100% protection against subsequent coinfection. The results indicate that when seasonal respiratory infections return to prepandemic levels, they could lead to an increased incidence of lethal COVID-19 superinfections, especially among the unvaccinated population.

Transient, inhaled gene therapy with gamma interferon mitigates pathology induced by host response in a mouse model of tuberculosis.

Transient transfection of the respiratory mucosa of mice infected with Mycobacterium tuberculosis (Mtb) with gamma interferon (IFN-γ) promises benefits in disease therapy. We investigated preclinical efficacy of a dry powder inhalation (DPI) as a stand-alone versus adjunct to oral anti-tuberculosis (TB) chemotherapy in mice. We observed that this host-directed therapy mitigates the gross organ pathology and histopathology of lung and spleen tissue of infected mice receiving the DPI, either alone or as adjunct therapy. However, no statistically significant reduction in Mtb colony forming units (CFU) occurred if mice were given only DPI; but not drugs. We compared one and three doses a week of the DPI over four weeks; with or without concomitant oral drugs. There was no significant difference in lung CFU after four or 12 doses of the DPI alone, but, surprisingly, four doses were qualitatively better than 12 doses in mitigating lung pathology. Nodular lesions on the lung surface and the area occupied by these was significantly reduced after four doses of the DPI, even without oral drugs. Transient transfection with IFN-γ did not induce pathological inflammation of the lungs and airways. We conclude that IFN-γ, as expected of host-directed therapy, 'heals the host; ' but does not 'kill the bug.'

Remodeling Yersinia pseudotuberculosis to generate a highly immunogenic outer membrane vesicle vaccine against pneumonic plague.

SignificanceYersinia pestis, the etiologic agent of plague, has been responsible for high mortality in several epidemics throughout human history. This plague bacillus has been used as a biological weapon during human history and is currently one of the deadliest biological threats. Currently, no licensed plague vaccines are available in the Western world. Since an array of immunogens are enclosed in outer membrane vesicles (OMVs), immune responses elicited by OMVs against a diverse range of antigens may reduce the likelihood of antigen circumvention. Therefore, self-adjuvanting OMVs from a remodeled Yersinia pseudotuberculosis strain as a type of plague vaccine could diversify prophylactic choices and solve current vaccine limitations.

Rubisco regulation in response to altered carbon status in the cyanobacterium Synechococcus elongatus PCC 7942.

Photosynthetic organisms possess a variety of mechanisms to achieve balance between absorbed light (source) and the capacity to metabolically utilize or dissipate this energy (sink). While regulatory processes that detect changes in metabolic status/balance are relatively well studied in plants, analogous pathways remain poorly characterized in photosynthetic microbes. Here, we explored systemic changes that result from alterations in carbon availability in the model cyanobacterium Synechococcus elongatus PCC 7942 by taking advantage of an engineered strain where influx/efflux of a central carbon metabolite, sucrose, can be regulated experimentally. We observed that induction of a high-flux sucrose export pathway leads to depletion of internal carbon storage pools (glycogen) and concurrent increases in estimates of photosynthetic activity. Further, a proteome-wide analysis and fluorescence reporter-based analysis revealed that upregulated factors following the activation of the metabolic sink are concentrated on ribulose-1,5-bisphosphate carboxylase-oxygenase (Rubisco) and auxiliary modules involved in Rubisco maturation. Carboxysome number and Rubisco activity also increased following engagement of sucrose secretion. Conversely, reversing the flux of sucrose by feeding exogenous sucrose through the heterologous transporter resulted in increased glycogen pools, decreased Rubisco abundance, and carboxysome reorganization. Our data suggest that Rubisco activity and organization are key variables connected to regulatory pathways involved in metabolic balancing in cyanobacteria.

Generation of Stable, Light-Driven Co-cultures of Cyanobacteria with Heterotrophic Microbes.

Co-cultivation of an autotrophic species with one or more heterotrophic microbes is a strategy for photobiological production of high-value compounds and is relatively underexplored in comparison to cyanobacterial or microalgal monocultures. Long-term stability of such consortia is required for useful collaboration between the partners, and this property can be increased by encapsulation of phototrophic partners within a hydrogel. Encapsulated cyanobacteria have advantages relative to planktonic cultures that may be useful to explore the potential for artificial microbial communities for targeted biomolecule synthesis, such as increased control over population sizes and reduced liquid handling requirements. In this chapter, we describe a method for encapsulation of genetically modified cyanobacterial strain (Synechococcus elongatus PCC 7942, CscB+) into a sodium alginate matrix, and the utilization of these encapsulated cells to construct stable, artificial autotroph/heterotroph co-cultures. This method has applications for the study of phototroph-based synthetic microbial consortia, and multi-species photobiological production.