Molecular basis for the increased activity of ZMS-2 serine protease in the presence of metal ions and hydrogen peroxide.

Serine proteases are important enzymes widely used in commercial products and industry. Recently, we identified a new serine protease from the desert bacterium Bacillus subtilis ZMS-2 that showed enhanced activity in the presence of Zn2+, Ag+, or H2O2. However, the molecular basis underlying this interesting property is unknown. Here, we report comparative studies between the ZMS-2 protease and its homolog, subtilisin E (SubE), from B. subtilis ATCC 6051. In the absence of Zn2+, Ag+, or H2O2, both enzymes showed the same level of proteolytic activity, but in the presence of Zn2+, Ag+, or H2O2, ZMS-2 displayed increased activity by 22%, 8%, and 14%, whereas SubE showed decreased activity by 16%, 12%, and 9%, respectively. In silico studies showed that both proteins have almost identical amino acid sequences and folding structures, except for two amino acids located in the protruding loops of the proteins. ZMS-2 contains Ser236 and Ser268, whereas SubE contains Thr236 and Thr268. Replacing Ser236 or Ser268 in ZMS-2 with threonine resulted in variants whose activities were not enhanced by Zn2+ or Ag+. However, this single mutation did not affect the enhancement by H2O2. This finding may be used as a basis for engineering better proteases for industrial uses.

Characterization and Pilot-Scale Application of the ZMS-2 Serine Protease with Novel Properties for the Eco-friendly Leather Processing.

Microbial alkaline proteases are dominating the global enzyme market with a share of over 65% due to their multifarious catalytic potentials. Hence, production of proteases with novel properties of commercial significance is highly desirable to meet the global enzyme demand. Here, we report the purification, characterization, and pilot-scale application of a serine protease from the desert soil bacterium Bacillus subtilis ZMS-2 with novel properties as dehairing agent in leather processing. The enzyme was purified 16.5-fold with a specific activity of 1543.5 U mg-1 and recovery percentage of 33.6% using ammonium sulfate precipitation, ion exchange, and gel filtration chromatography. The purified enzyme was characterized as a metal ion-, surfactant-, and denaturant-compatible alkaline serine protease having a molecular weight of 36.1 kDa with an optimum activity at pH 8.5 and 60 °C. The catalytic activity of the enzyme was enhanced by Zn+2 (204%), Ag+ (110%), H2O2 (123%), Triton X-100 (110%), iso-octane (109%), chloroform (110%), ethanol (105%), ethyl acetate (110%), and acetonitrile (128%). During pilot-scale applications, the optimum condition was found to be a combination of enzyme (1.5%, 460 U mL-1), sodium sulfide (2%), and calcium hydroxide (lime) (3%). Under this condition, the time required for complete dehairing was 90 min. Chemoenzymatically processed skins exhibited better physical properties than chemically processed skin, including tensile strength (16.35 ± 6.68 N/mm), ball burst (452.88 ± 6.06 N/mm), percent elongation (38.85 ± 1.06 N), tear strength (50.16 ± 4.42 N/mm), and softness (6.5 mm). Electron microscopy analysis of the treated skin showed complete removal of hairs with roots, confirming the keratin specificity of the enzyme. Moreover, the enzyme-assisted dehairing process reduced chemical oxygen demand (COD), biochemical oxygen demand (BOD), total dissolved solids (TDS), and total suspended solids (TSS) by 68, 77, 34, and 39%, respectively. Thus, the alkaline serine protease from B. subtilis ZMS-2 is a potential dehairing agent for the eco-friendly processing of animal skins on industrial scales.

In-vitro evaluation of virulence markers and antifungal resistance of clinical Candida albicans strains isolated from Karachi, Pakistan.

Candidiasis is a significant fungal infection with high mortality and morbidity rates worldwide. Candida albicans is the most dominant species responsible for causing different manifestations of candidiasis. Certain virulence traits as well as its resistance to antifungal drugs contribute to the pathogenesis of this yeast. This study was designed to determine the production of some virulence factors, such as biofilm formation and extracellular hydrolytic enzymes (esterase, coagulase, gelatinase, and catalase) by this fungus, as well as its antifungal resistance profile. A total of 304 clinical C. albicans isolates obtained from different clinical specimens were identified by a conventional diagnostic protocol. The antifungal susceptibility of C. albicans strains was determined by disk diffusion technique against commercially available antifungal disks, such as nystatin 50 µg, amphotericin B 100 unit, fluconazole 25 µg, itraconazole 10 µg, ketoconazole 10 µg, and voriconazole 1 µg. The assessment of biofilm formation was determined by the tube staining assay and spectrophotometry. Gelatinase, coagulase, catalase, and esterase enzyme production was also detected using standard techniques. A total of 66.1% (201/304) and 28.9% (88/304) of C. albicans strains were susceptible-dose dependent (SDD) to nystatin and itraconazole, respectively. Among the antifungal drugs, C. albicans strains showed high resistance to ketoconazole 24.7% (75/304); however, no statistically significant relationship between the clinical origin of C. albicans isolates and antifungal drug resistance pattern was detected. For virulence factors, the majority of the C. albicans strains actively produced biofilm and all hydrolytic enzymes. Biofilm formation was demonstrated by 88% (267/304) of the strains with a quantitative mean value 0.1762 (SD ± 0.08293). However, 100% (304/304) of isolates produced catalase enzyme, 69% (211/304) produced coagulase, 66% (197/304) produced gelatinase, and 52% (157/304) produced esterase enzyme. A significant relationship between the source of specimens and biofilm formation by C. albicans was observed; nevertheless, there was no significant relationship between different sources of C. albicans strains and the production of different enzymatic virulence factors. The study found that C. albicans strains have excellent potential to produce virulence markers and resistance to antifungals, which necessitates surveillance of these opportunistic pathogens to minimize the chances of severe invasive infections.

Design and Immunoinformatic Assessment of Candidate Multivariant mRNA Vaccine Construct against Immune Escape Variants of SARS-CoV-2.

To effectively counter the evolving threat of SARS-CoV-2 variants, modifications and/or redesigning of mRNA vaccine construct are essentially required. Herein, the design and immunoinformatic assessment of a candidate novel mRNA vaccine construct, DOW-21, are discussed. Briefly, immunologically important domains, N-terminal domain (NTD) and receptor binding domain (RBD), of the spike protein of SARS-CoV-2 variants of concern (VOCs) and variants of interest (VOIs) were assessed for sequence, structure, and epitope variations. Based on the assessment, a novel hypothetical NTD (h-NTD) and RBD (h-RBD) were designed to hold all overlapping immune escape variations. The construct sequence was then developed, where h-NTD and h-RBD were intervened by 10-mer gly-ala repeat and the terminals were flanked by regulatory sequences for better intracellular transportation and expression of the coding regions. The protein encoded by the construct holds structural attributes (RMSD NTD: 0.42 Å; RMSD RBD: 0.15 Å) found in the respective domains of SARS-CoV-2 immune escape variants. In addition, it provides coverage to the immunogenic sites of the respective domains found in SARS-CoV-2 variants. Later, the nucleotide sequence of the construct was optimized for GC ratio (56%) and microRNA binding sites to ensure smooth translation. Post-injection antibody titer was also predicted (~12000 AU) to be robust. In summary, the construct proposed in this study could potentially provide broad spectrum coverage in relation to SARS-CoV-2 immune escape variants.

Characterization of the genome and serine protease of a novel Bacillus subtilis isolate.

Current study was undertaken to carry out the genome-wide analysis of a multipotent isolate from desert soil which was previously identified as Bacillus tequilensis based on 16S rDNA analysis. This study also aims to characterize the serine protease and its biocatalytic potentials implying a combination of empirical and in-silico approaches. Next generation sequencing and short read de novo assembly generated the 4,235,084 bp draft genome of Bacillus sp. ZMS-2. Genome sequence analysis by digital DNA:DNA hybridization (dDDH) and average nucleotide identity classified the isolate as Bacillus subtilis ZMS-2 (Bioproject ID: PRJNA691551). Genome annotation revealed 10 antibiotic resistance genes, 8 antibiotic/antifungal gene clusters and 25 genes encoding proteases including subtilisin E, an extracellular alkaline protease. This extracellular protease (ZMS-2 protease) was produced using a statistically optimized medium, purified partially and characterized as alkaline serine protease. The partially purified ZMS-2 protease (780 U/mL) showed a 21 mm zone of casein hydrolysis and dehaired goat skin by pulling out hair with roots. These catalytic potentials of ZMS-2 protease were further confirmed using scanning electron microscopy of casein beads and dehaired skin. The study concludes B. subtilis ZMS-2 as a potent producer of a protease with promising potentials of commercial importance.

Public Perceptions of Current COVID-19 Vaccinations. Results of a Pilot Survey.

INTRODUCTION: We conducted a cross-sectional survey as a part of an educational program in collaboration with the Global Thrombosis Forum (GTF), an affiliate of North American Thrombosis Forum (NATF), and Loyola University about public perceptions of COVID-19 and COVID-19 vaccinations in the US. In this study, we are reporting the results of this survey. MATERIALS AND METHODS: The survey, in the form of a questionnaire, has been developed by GTF and faculty members. A prepared questionnaire was sent to the members of the Georgia and Illinois communities. RESULTS: In our current study, the COVID-19 vaccine willingness rate was 94.5% and vaccination rate was 90.9%. In multivariate analysis believing to have enough information about the safety and efficacy of COVID-19 vaccines (OR: 3.730, 95% CI: 1.199-11.603, p: 0.023) and gender (OR: 0.123, 95% CI: 0.016-0.967, p: 0.046) were significant predictors for vaccine willingness. Previous COVID-19 infection (OR: 0.215, 95% CI: 0.061-0.758, p: 0.017), moderate and severe effects of COVID-19 pandemic on participant's life (OR: 4.631, 95% CI 1.681-12.760, p: 0.003) and believing to have enough information about the safety and efficacy of COVID-19 vaccines (OR: 4.119, 95% CI: 1.508-11.253, p: 0.006) were significant predictors for final vaccination status. CONCLUSION: In conclusion, currently vaccination remains one of the most effective tools in the fight against the COVID-19 pandemic. The vaccine hesitancy is a complex phenomenon that is driven by individuals' perceptions of safety, and efficiency of the vaccines. We must continue to educate the public and communities that vaccines are safe, that they are effective and that they are still required even after a COVID-19 infection.

The COVID-19 Pandemic and the Need for an Integrated and Equitable Approach: An International Expert Consensus Paper.

BACKGROUND: One year after the declaration of the coronavirus disease 2019 (COVID-19) pandemic by the World Health Organization (WHO) and despite the implementation of mandatory physical barriers and social distancing, humanity remains challenged by a long-lasting and devastating public health crisis. MANAGEMENT: Non-pharmacological interventions (NPIs) are efficient mitigation strategies. The success of these NPIs is dependent on the approval and commitment of the population. The launch of a mass vaccination program in many countries in late December 2020 with mRNA vaccines, adenovirus-based vaccines, and inactivated virus vaccines has generated hope for the end of the pandemic. CURRENT ISSUES: The continuous appearance of new pathogenic viral strains and the ability of vaccines to prevent infection and transmission raise important concerns as we try to achieve community immunity against severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) and its variants. The need of a second and even third generation of vaccines has already been acknowledged by the WHO and governments. PERSPECTIVES: There is a critical and urgent need for a balanced and integrated strategy for the management of the COVID-19 outbreaks organized on three axes: (1) Prevention of the SARS-CoV-2 infection, (2) Detection and early diagnosis of patients at risk of disease worsening, and (3) Anticipation of medical care (PDA). CONCLUSION: The "PDA strategy" integrated into state policy for the support and expansion of health systems and introduction of digital organizations (i.e., telemedicine, e-Health, artificial intelligence, and machine-learning technology) is of major importance for the preservation of citizens' health and life world-wide.

Molecular docking between human TMPRSS2 and SARS-CoV-2 spike protein: conformation and intermolecular interactions.

Entry of SARS-CoV-2, etiological agent of COVID-19, in the host cell is driven by the interaction of its spike protein with human ACE2 receptor and a serine protease, TMPRSS2. Although complex between SARS-CoV-2 spike protein and ACE2 has been structurally resolved, the molecular details of the SARS-CoV-2 and TMPRSS2 complex are still elusive. TMPRSS2 is responsible for priming of the viral spike protein that entails cleavage of the spike protein at two potential sites, Arg685/Ser686 and Arg815/Ser816. The present study aims to investigate the conformational attributes of the molecular complex between TMPRSS2 and SARS-CoV-2 spike protein, in order to discern the finer details of the priming of viral spike protein. Briefly, full length structural model of TMPRSS2 was developed and docked against the resolved structure of SARS-CoV-2 spike protein with directional restraints of both cleavage sites. The docking simulations showed that TMPRSS2 interacts with the two different loops of SARS-CoV-2 spike protein, each containing different cleavage sites. Key functional residues of TMPRSS2 (His296, Ser441 and Ser460) were found to interact with immediate flanking residues of cleavage sites of SARS-CoV-2 spike protein. Compared to the N-terminal cleavage site (Arg685/Ser686), TMPRSS2 region that interact with C-terminal cleavage site (Arg815/Ser816) of the SARS-CoV-2 spike protein was predicted as relatively more druggable. In summary, the present study provides structural characteristics of molecular complex between human TMPRSS2 and SARS-CoV-2 spike protein and points to the candidate drug targets that could further be exploited to direct structure base drug designing.

Dataset for homologous proteins in Drosophila melanogaster for SARS-CoV-2/human interactome.

Animal modelling for infectious diseases is critical to understand the biology of the pathogens including viruses and to develop therapeutic strategies against it. Herein, we present the sequence homology and expression data analysis of proteins found in Drosophila melanogaster that are orthologous to human proteins, reported as components of SARS-CoV-2/Human interactome. The dataset enlists sequence homology, query coverage, domain conservation, OrthoMCL and Ensembl Genome Browser support of 326 proteins in D.melanogaster that are potentially orthologous to 417 human proteins reported for their direct physical interactions with 28 proteins encoded by SARS-CoV-2 genome. Expression of these D.melanogaster orthologous genes in 26 anatomical positions are also plotted as heat maps in 27 sets, corresponding to the potential protein interactors for each viral protein. The data could be used to direct experiments and potentially predict their phenotypic and molecular outcome in order to dissect the biological roles and molecular functionality of SARS-CoV-2 proteins in a convenient animal model system like D.melanogaster.

Luffa cylindrica Immobilized with Aspergillus terreus QMS-1: an Efficient and Cost-Effective Strategy for the Removal of Congo Red using Stirred Tank Reactor.

Microbial populations within the rhizosphere have been considered as prosperous repositories with respect to bioremediation aptitude. Among various environmental contaminants, effluent from textile industries holds a huge amount of noxious colored materials having high chemical oxygen demand concentrations causing ecological disturbances. The study was aimed to explore the promising mycobiome of rhizospheric soil for the degradation of azo dyes to develop an efficient system for the exclusion of toxic recalcitrants. An effluent sample from the textile industry and soil samples from the rhizospheric region of Musa acuminata and Azadirachta indica were screened for indigenous fungi to decolorize Congo red, a carcinogenic diazo dye, particularly known for its health hazards to the community. To develop a bio-treatment process, Aspergillus terreus QMS-1 was immobilized on pieces of Luffa cylindrica and exploited in stirred tank bioreactor under aerobic and optimized environment. Quantitative estimation of Congo red decolorization was carried out using UV-Visible spectrophotometer. The effects of fungal immobilization and biosorption on the native structure of Luffa cylindrica were evaluated using a scanning electron microscope. A. terreus QMS-1 can remove (92%) of the dye at 100 ppm within 24 h in the presence of 1% glucose and 1% ammonium sulphate at pH 5.0. The operation of the bioreactor in a continuous flow for 12 h with 100 ppm of Congo red dye in simulated textile effluent resulted in 97% decolorization. The stirred tank bioreactor was found to be a dynamic, well maintained, no sludge producing approach for the treatment of textile effluents by A. terreus QMS-1 of the significant potential for decolorization of Congo red.Microbial populations within the rhizosphere have been considered as prosperous repositories with respect to bioremediation aptitude. Among various environmental contaminants, effluent from textile industries holds a huge amount of noxious colored materials having high chemical oxygen demand concentrations causing ecological disturbances. The study was aimed to explore the promising mycobiome of rhizospheric soil for the degradation of azo dyes to develop an efficient system for the exclusion of toxic recalcitrants. An effluent sample from the textile industry and soil samples from the rhizospheric region of Musa acuminata and Azadirachta indica were screened for indigenous fungi to decolorize Congo red, a carcinogenic diazo dye, particularly known for its health hazards to the community. To develop a bio-treatment process, Aspergillus terreus QMS-1 was immobilized on pieces of Luffa cylindrica and exploited in stirred tank bioreactor under aerobic and optimized environment. Quantitative estimation of Congo red decolorization was carried out using UV-Visible spectrophotometer. The effects of fungal immobilization and biosorption on the native structure of Luffa cylindrica were evaluated using a scanning electron microscope. A. terreus QMS-1 can remove (92%) of the dye at 100 ppm within 24 h in the presence of 1% glucose and 1% ammonium sulphate at pH 5.0. The operation of the bioreactor in a continuous flow for 12 h with 100 ppm of Congo red dye in simulated textile effluent resulted in 97% decolorization. The stirred tank bioreactor was found to be a dynamic, well maintained, no sludge producing approach for the treatment of textile effluents by A. terreus QMS-1 of the significant potential for decolorization of Congo red.

Structural variations in human ACE2 may influence its binding with SARS-CoV-2 spike protein.

The recent pandemic of COVID-19, caused by SARS-CoV-2, is unarguably the most fearsome compared with the earlier outbreaks caused by other coronaviruses, SARS-CoV and MERS-CoV. Human ACE2 is now established as a receptor for the SARS-CoV-2 spike protein. Where variations in the viral spike protein, in turn, lead to the cross-species transmission of the virus, genetic variations in the host receptor ACE2 may also contribute to the susceptibility and/or resistance against the viral infection. This study aims to explore the binding of the proteins encoded by different human ACE2 allelic variants with SARS-CoV-2 spike protein. Briefly, coding variants of ACE2 corresponding to the reported binding sites for its attachment with coronavirus spike protein were selected and molecular models of these variants were constructed by homology modeling. The models were then superimposed over the native ACE2 and ACE2-spike protein complex, to observe structural changes in the ACE2 variants and their intermolecular interactions with SARS-CoV-2 spike protein, respectively. Despite strong overall structural similarities, the spatial orientation of the key interacting residues varies in the ACE2 variants compared with the wild-type molecule. Most ACE2 variants showed a similar binding affinity for SARS-CoV-2 spike protein as observed in the complex structure of wild-type ACE2 and SARS-CoV-2 spike protein. However, ACE2 alleles, rs73635825 (S19P) and rs143936283 (E329G) showed noticeable variations in their intermolecular interactions with the viral spike protein. In summary, our data provide a structural basis of potential resistance against SARS-CoV-2 infection driven by ACE2 allelic variants.

Prevalence and association of HBV and HCV infection with cardiovascular disease risk factors in a peri-urban population.

OBJECTIVE: To investigate the possible associations of hepatitis B and C virus infection with cardiovascular disease risk factors inperi-urban population. METHODS: The cross-sectional study was conducted from February to December 2016 in the periurban low-resource locality of Bin Qasim Town in Karachi. Serum samples were screened for hepatitis B surface antigen and hepatitis C virus antibodies. Anthropometric measurements were taken and markers related to cardiovascular disease were examined. Association of the two hepatitis virus infections with cardiovascular diseasewere investigated by anaylsing the data using SPSS 16. RESULTS: There were 691 subjects. Serum triglyceride levels were significantly low in patients with hepatitis B virus (p<0.05). Those with hepatitis C virus had markedly low total cholesterol, low-density lipoprotein cholesterol, high-density lipoprotein cholesterol and triglyceride levels (p<0.05 each), whereas random blood sugar and body mass index values were significantly high (p<0.05 each. Hepatitis C virus infection was positively associated with body mass index and random blood glucose, and inversely associated with total cholesterol, high-density lipoprotein cholesterol and low-density lipoprotein cholesterol(p< 0.05 each). CONCLUSIONS: Hepatitis B virus infection showed a significant inverse association with triglyceride levels. However, hepatitis C virus infection was positively associated with body mass index and random blood sugar, and inversely associated with total cholesterol, high-density lipoprotein cholesterol and low-density lipoprotein cholesterol levels.

Bacillus tequilensis ZMS-2: A novel source of alkaline protease with antimicrobial, anti-coagulant, fibrinolytic and dehairing potentials.

Emerging resistance to existing antimicrobial agents is one of the growing concerns and a serious problem for public health globally. Currently available antimicrobial agents are potent and effective but surfacing resistance to these drugs has not been ruled out so far. Therefore, it is utmost important to explore new bioactive compounds from natural sources to meet future needs. The present study was designed to produce, optimize, characterize and evaluate antimicrobial, fibrinolytic and anti-coagulant potential of a new alkaline protease. Proteolytic strain from desert soils of Tharparkar, Pakistan was subjected to 16S rDNA sequencing and identified as Bacillus tequilensis ZMS-2(Genbank Accession No. MK101013). During submerged fermentation at 37ºC, maximum enzyme production (454 U/ml) was observed with 24h old inoculum. The best incubation time was 72h (544 U/ml), optimum inoculum size and pH was 10% at pH 8 with 494 and 506 U/ml, respectively. The best carbon source was starch (571 U/ml), while ideal substrate was wheat bran (536 U/ml). Optimal temperature and pH for proteolytic activity was 60ºC (420 U/ml) and 8 (332 U/ml). Alkaline protease showed antibacterial activity against Staphylococcus aureus (27mm), Bacillus licheniformis (20mm), Klebsiella pneumoniae (17mm) and Escherichia coli (15mm). The strain B. tequilensis ZMS-2 also exhibited anticoagulant, fibrinolytic and dehairing potential suggesting application of its protease in various industries.

Non-albicans Candida species: Emergence of neglected pathogens among population of Karachi.

Candida albicans was considered as the principal cause of opportunistic candidiasis but nowadays, neglected non-albicans Candida (NAC) species are evolving as more virulent and drug resistant strains. This research was intended to assess pervasiveness of candidiasis mainly caused by NAC species in Karachi city. A total of 562 clinical isolates of Candida spp. collected during the period of one year were identified by microscopic as well as morphological (germ tube formation, characteristics on CHROM agar and Corn meal agar) and Biochemical (sugar assimilation and fermentation) characteristics. Doubtful species were further identified by using Remel RapIDTM yeast plus kit. The results were statistically analyzed by SPSS 16.0 version software. Isolated strains of candida revealed slight predominance of C. albicans (54.5%) over non- albicans Candida species (45.5%). Among NAC species, C. tropicalis and C. glabrata were isolated as the predominant species. These clinical species were procured mainly from urine samples of females (73.7%) of age group 20-30 years. No significant correlations exist between Candida species and their months of isolation as well as their isolation from different districts of Karachi. Emergence of NAC species may predict an upcoming threat in health care facilities and hence, require prompt management and accurate identification to suggest empirical antifungal therapy.

Antimicrobial potential of newly isolated Aspergillusterreus MK-1: An approach towards new antibiotics.

OBJECTIVE: To attempt discovering new bioactive metabolites from fungal sources. METHODS: The exploratory study was conducted at the Department of Microbiology, Federal Urdu University for Arts, Science and Technology, Karachi from January 2016 to November 2017and comprised of soil samples collected from rhizosphere region of different garden plants from the city. Fungi were screened for production of antibiotics by testing cell-free culture filtrates obtained by Shake-flask fermentation technique. Agar-Well diffusion assay method was used to evaluate antagonistic activity against pathogenic microorganisms. RESULTS: Bioactive compounds extracted by ethyl acetate and thin layer chromatography revealed mixture of compounds in the crude extract. AspergillusterreusMK-1 showed significant inhibition of medically important test pathogens namely Staphylococcus aureus, Pseudomonas aeruginosa, Escherichiacoli, Salmonella typhi, Micrococcus luteus, Streptococcus epidermidis, Bacillus subtilis, Candida albicans and Aspergillusniger. The best biological activity of crude ethyl acetate extract was observed against Pseudomonas aeruginosa (63mm). CONCLUSIONS: Newly isolated AspergillusterreusMK-1 emerged as a potent candidate for the production of antimicrobial compounds.

Fungal contamination in smokeless tobacco products traditionally consumed in Pakistan.

OBJECTIVE: To isolate potential pathogenic fungi from smokeless tobacco products. METHODS: The study was conducted from January 2015 to February 2017 during which samples of smokeless tobacco products such as Mainpuri, Tambako, Khiwam, Gutkha, Naswar and Mawa etc. were collected from different cities of Pakistan. The samples were tested for fungal contamination by spread plate method. Different strains of fungi were isolated and identified on the basis of their macroscopic as well as microscopic characteristics. The fungal strains isolated were also screened for their susceptibility to commonly used antifungal drugs by disc diffusion method. RESULTS: Of the 600 samples collected, 300(50%) were from Sindh, 70(11.7%) Balochistan, 74(12.3%) from Khyber Pakhtunkhwa, 105(17.5%) from Punjab and 51(8.5%) from Azad Kashmir. In terms of products, there were 404(67.3%) samples of Naswar, 69(11.5%) Patti, 40(6.6%) Khiwam, 35(5.8%) Mawa, 32(5.3%) Gutkha, and 20(3.3%) Mainpuri samples. Different species of Aspergillus were predominantly isolated followed by Penicillium, Mucor, Sepedonium and Trichophyton. The isolated strains of Aspergillus also revealed resistance against many commonly-used anti-fungals such as Amphotericin B and Itraconazole.. CONCLUSIONS: There was high prevalence of opportunistic fungi in study samples, posing a threat for human health which requires prompt notice and management.

The Pattern of Psychiatric Morbidity in an Outpatient Child Psychiatry Clinic: A Cross-sectional, Descriptive Study from a Tertiary Care Hospital in Kashmir, North India.

BACKGROUND: Psychiatric disorders are ubiquitous and affect not only adults but also children and adolescents. The age factor plays an important role in the pattern of these psychiatric disorders. The objective of our study was to find the pattern of psychiatric morbidity in children and adolescents at the child and adolescent outpatient service of a tertiary care hospital. MATERIALS AND METHODS: A semi-structured questionnaire was used to record the sociodemographic status. The state of mental health and psychiatric morbidity was assessed after a thorough clinical assessment. Intelligence quotient was assessed by a clinical psychologist as and when needed. All the diagnoses were made on the basis of Diagnostic and Statistical Manual of Mental Disorders, 4th Edition, Text Revision criteria. RESULTS: A total of 529 patients were included. Most patients belonged to the age group of 6-16 years (70.5%). Boys (67.9%) outnumbered girls. Most of the patients were from rural background (56.7%) and from nuclear families (53%). Attention-deficit hyperactivity disorder (31%) and mental retardation (29%) were the most frequent diagnoses, followed by pervasive developmental disorders (10%). Comorbidity was present in about 18% of our patients. CONCLUSION: The child psychiatry is gaining acceptance, and children and adolescents with minor mental health issues are being identified and referred for specialized services.

Isolation of potentially pathogenic fungi from selected pigeons' feeding sites in Karachi: A new dimension to health hazard.

OBJECTIVE: To determine the presence of pathogenic fungal strains in areas where pigeons are present in a large number. METHODS: This study was conducted at the Federal Urdu University of Arts, Science and Technology, Karachi, from February 2015 to March2016, and comprised samples of soil contaminated with pigeons' excreta. The samples were collected from 20 different pigeon-feeding places in the city. These samples were processed for the isolation and identification of fungi by using standard conventional methods. The fungal strains isolated were also tested for their susceptibility to commonly used antifungal agents by disc diffusion technique. RESULTS: There were 105 samples. A wide variety of fungal strains belonging to different genera of Aspergillus, Rhizopus, Penicillium, Fusarium and Candida were isolated and identified by using conventional methods. The antifungal resistance pattern of these strains also depicts emergence of resistance against commonly used antifungal agents such as amphotericin B and fluconazole. CONCLUSIONS: The soil and air of places densely populated with pigeons were found to be loaded with fungal spores and many of them were potential pathogens.

Biophysicochemical characterization of Pyocin SA189 produced by Pseudomonas aeruginosa SA189.

Pseudomonas aeruginosa, in spite of being a ubiquitous organism (as it is found in soil, water, and humans), is also an opportunistic pathogen. In order to maintain its diversity in the community, it produces various toxic proteins, known as, bacteriocins. In the present study, pyocin SA189, which is a bacteriocin produced by P. aeruginosa SA189 (isolated from a clinical sample) was characterized. P. aeruginosa SA189, as identified by the conventional and 16S rRNA gene amplification, produced pyocin SA189 of molecular weight of 66 k Da. The pyocin showed antimicrobial activity against several clinically relevant Gram-positive and Gram-negative bacteria and was substantially stable for wide ranges of temperature and pH. Furthermore, the pyocin also retained its biological activity upon treatment with metal ions, organic solvents, and various proteolytic and lipolytic enzymes. The data from the growth kinetics indicated that the maximum bacteriocin production occurred in the late log phase. Overall, our results signify the potential of pyocin SA189 as a bio-control agent.

Biophysicochemical characterization of Pyocin SA189 produced by Pseudomonas aeruginosa SA189

Abstract Pseudomonas aeruginosa, in spite of being a ubiquitous organism (as it is found in soil, water, and humans), is also an opportunistic pathogen. In order to maintain its diversity in the community, it produces various toxic proteins, known as, bacteriocins. In the present study, pyocin SA189, which is a bacteriocin produced by P. aeruginosa SA189 (isolated from a clinical sample) was characterized. P. aeruginosa SA189, as identified by the conventional and 16S rRNA gene amplification, produced pyocin SA189 of molecular weight of 66 k Da. The pyocin showed antimicrobial activity against several clinically relevant Gram-positive and Gram-negative bacteria and was substantially stable for wide ranges of temperature and pH. Furthermore, the pyocin also retained its biological activity upon treatment with metal ions, organic solvents, and various proteolytic and lipolytic enzymes. The data from the growth kinetics indicated that the maximum bacteriocin production occurred in the late log phase. Overall, our results signify the potential of pyocin SA189 as a bio-control agent.