Alleviation of cadmium toxicity and growth enhancement of helianthusannuus and triticum aestivum seedlings through bacterial inoculation

Two Cd-resistant bacterial growth strains MA-9 (Aeromonas) and MA19 (affinities uncertain), which were isolated from ICI effluents, were used to inoculate seeds ifTriticum aestivum (wheat) and Helianthus annuus (sunflower). Both inoculated and non-inoculated seeds were germinated and grown under different concentrations (0, 1, 2 and 3 mM) of CdCl2 for 10 days. Under Cd-stress conditions, bacteria-inoculated plants had better germination and growth than non-inoculated treatments. Bacterial growth enhancement if seedlings was associated with reduced Cd uptake

Evaluation of bla SHV, bla TEM and bla OXA encoding Clinical Isolates from Chronic Tonsillitis using Phenotypic and Molecular Technique: First report from Pakistan

Objective: To evaluate bla SHV, bla TEM and bla OXA encoding clinical isolates in chronic tonsillitis using phenotypic and molecular techniques.Place and Duration:  The study was conducted in Department of Microbiology and Molecular Genetics, University of Punjab, Lahore from August 2016 to August 2017.Methodology: Sample processing, identification and characterization of isolates was done by using (CLSI, 2016) criteria. Antibiotic susceptibility testing by using disc diffusion assay and biofilm forming ability was analyzed by ring test and slime production test. Combination disc test was used for phenotypic detection of antibiotic resistance genes. Multiplex-PCR assay was used to check the presence of bla SHV, bla TEM and bla OXA genes. 16S rRNA sequencing and phylogenetic analysis was performed.Results: Here, variable resistance pattern was observed against applied antibiotics. 100 % resistance towards azotreonam and penicillin was observed. While 60-85 % resistances were observed against cephalosporins. Biofilm formation increased with the passage of time. 77% strains indicated positive combination disc test. Multiplex-PCR indicated 60% strains harbored tested genes. 40 % bla SHV genes, 30 % bla TEM genes and 60% bla OXA genes were observed among selected isolates. GenBank Accession number obtained for Klebsiella pneumoniae was KY810693 and for S. aureus was KY810692.Conclusion: In conclusion, K. pneumoniae and S. aureus came out to be common causative agents of tonsillitis in the current study. Resistance towards multiple classes of antibiotics and strong biofilms of these micro-organisms explain the chronicity and recurrent nature of the infection. bla OXA genes were frequent among genes tested

Bacterial exopolysaccharide and biofilm formation stimulate chickpea growth and soil aggregation under salt stress

To compensate for stress imposed by salinity, biofilm formation and exopolysaccharide production are significant strategies of salt tolerant bacteria to assist metabolism. We hypothesized that two previously isolated salt-tolerant strains Halomonas variabilis (HT1) and Planococcus rifietoensis (RT4) have an ability to improve plant growth, These strains can form biofilm and accumulate exopolysacharides at increasing salt stress. These results showed that bacteria might be involved in developing microbial communities under salt stress and helpful in colonizing of bacterial strains to plant roots and soil particles. Eventually, it can add to the plant growth and soil structure. We investigated the comparative effect of exopolysacharide and biofilm formation in two bacterial strains Halomonas variabilis (HT1) and Planococcus rifietoensis (RT4) in response to varying salt stress. We found that biofilm formation and exopolysaccharide accumulation increased at higher salinity. To check the effect of bacterial inoculation on the plant (Cicer arietinum Var. CM-98) growth and soil aggregation, pot experiment was conducted by growing seedlings under salt stress. Inoculation of both strains increased plant growth at elevated salt stress. Weight of soil aggregates attached with roots and present in soil were added at higher salt concentrations compared to untreated controls. Soil aggregation was higher at plant roots under salinity. These results suggest the feasibility of using above strains in improving plant growth and soil fertility under salinity

Computational and experimental strategies for combating MBL P. aeruginosa (MBLPA) biofilms using phytochemicals: Targeting the quorum sensing network

Pseudomonas aeruginosa is a Gram-negative opportunistic bacterium, ubiquitously found in nature and causative agent in many infections. Due to increased antibiotic resistance, there is a need to develop more robust antibacterial agents from natural sources. In this study, we worked on two metallo-β-lactamase (MBL) producing Pseudomonas aeruginosa strains and targeted the Quorum Sensing mechanism (QS) of these bacteria to combat antibiotic resistance. Our study aimed at using phytochemicals which have been used since centuries in herbal medicine. We used fifteen commercially available phytochemicals and check their effects on biofilm formation, quorum sensing and inter-related mechanisms. Sub-inhibitory concentration of isoliquiritin inhibited biofilm formation 55 % in P8 at day 6 and 48 % in P6 at day 6; quorum sensing 83 % in P6 and 61 % in P8 whereas sub-inhibitory concentration of 6-gingerol suppressed biofilm formation by 48 % in P8 at day 6 and 44 % in P6 at day 6; quorum sensing 69 % in P6 and 48 % in P8, respectively. The results indicated isoliquiritin, epigallocatechin gallate, eugenol, luteolin and chrysin to be the potential candidates in inhibiting QS and related mechanisms. Isoliquiritin which was never been used before against biofilm and QS related studies, showed remarkable results and found to be more efficient in inhibiting QS than 6-gingerol –a known QS inhibitor. For examining the molecular interaction between phytochemicals and QS, In-silico molecular docking was performed between phytoligands and four QS proteins (Las I, Las R, RhlI and Rhl R). In-silico docking analysis revealed that isoliquiritin showed strong bond with amino acids (Trp34, Asp35, Asp35, Tyr105, Arg104, Val138, Thr140) present at the active site of RhlI with binding energy value of −8.4 kcal/mol as compared to that of 6-gingerol with Rhl1 (−7.3 kcal/mol). In conclusion, our study may help in controlling nosocomial infections caused by carbapenem-resistant metallo beta-lactamase P. aeruginosa (MBL-PA) by utilizing these phytochemicals in biofilms disruption and quorum sensing inhibition. Moreover their synergism with antibiotics may help in lowering the MIC of carbapenem antibiotics against such Multi-drug resistant strains

HLA Association in SLE patients from Lahore-Pakistan

The first genetic factors to be identified as important in the pathogenesis of Systemic lupus erythematosus (SLE) were those of the major histocompatibility complex (MHC) on chromosome 6. It is now widely accepted that MHC genes constitute a part of the genetic susceptibility to SLE. The study population comprised 61 SLE patients fulfilling at least four of the American college of Rheumatology criteria for SLE and 61 healthy blood donors as controls. SLE female versus male ratio was approximately 9:1. Mean age at diagnosis was 30.35 ± 1.687 (12-68 years). DNA-based HLA Typing for HLA-A, HLA-B, and HLA-DRB1 was carried out by Polymerase chain reaction with sequence specific primers using genomic DNA obtained from blood samples. A total of 22 alleles have been studied at locus A, 37 alleles at locus B and 17 DRB1 alleles. The allelic frequencies of HLA-A, HLA-B, and HLA-DRB1 antigens in SLE patients from Pakistan were compared with the controls. A significant increase was observed in the frequency of HLA-A*01, A*03, A*11, A*23, A*26 A*69, HLA-B*27, B*40, B*49, B*51, B*52, B*53, B*54, B*95, HLA-DRBI*01, DRBI*03, DRBI*11, DRBI*14 among SLE patients indicating a positive association of these alleles with SLE. HLA-A*24, A*29, A*31, A*34, A*68, A*92, HLA-B*18, HLA-DRB1*12, were found to be decreased in the patient group as compared to controls indicating a negative association of these alleles with SLE. Thus from this study we can conclude that SLE is associated with certain MHC alleles in Pakistani population

Angiotensin-converting enzyme gene I/D polymorphism in Pakistani systemic lupus erythematosus patients

Angiotensin-converting enzyme (ACE) was first identified as a key component of the rennin-angiotensin system, as its main role is to process angiotensin I to angiotensin II and degrade bradykinin. Human ACE maps to chromosome 17q23 spans 21Kb, includes 26 exons and 25 introns. In humans, ID, DD, and II  polymorphism is located in intron 16 of the angiotensin gene. The purpose of this study is to investigate the frequency of ACE gene insertion/deletion (I/D)  polymorphism genotype in systemic lupus  erythematosus (SLE) patients and to study the correlation between I/D polymorphism of the ACE gene and clinical  manifestations of SLE. Sixty one (61) controls and 61 SLE patients were recruited from Punjab-Pakistan. Sixty one SLE patients and 61 control subjects were  studied for ACE I/D  polymorphism by using Triple primer method with nested polymerase chain reaction (PCR). The frequency of DD, ID and II genotypes was 54, 3 and 4% in SLE patients’ and 23, 32 and 6% in healthy controls, respectively. The frequency of DD allele in SLE patients with lupus nephritis is 100%, Sjogren’s syndrome 100%, Raynaud’s phenomenon 88.88%, and with rheumatoid arthritis it is 78.94%. The frequency of ID allele in SLE patients with Raynaud’s  phenomenon is 5.55%, and with rheumatoid arthritis it is 10.52%. The frequency of II allele in SLE patients with Raynaud’s phenomenon 5.55%, rheumatoid  arthritis is 10.52% but the important thing to note is that the frequency of II allele in SLE patients with vasculitis is 100%. This study was undertaken to determine whether DD, ID and II polymorphisms of Intron16 of the ACE gene is associated with SLE and whether the results support such an association. It can be concluded that lupus nephritis, Sjogren’s syndrome, Raynaud’s phenomenon, rheumatoid arthritis and vasculitis, which are common among Pakistani SLE patients, are  related diseases and ACE gene is involved in lupus  susceptibility.Key words: Systemic lupus erythematosus, angiotensin converting enzyme I/D polymorphism,  Sjogren’s syndrome, Raynaud’s phenomenon, rheumatoid  arthritis, vasculitis

Evaluation of bla SHV, bla TEM and bla OXA encoding Clinical Isolates from Chronic Tonsillitis using Phenotypic and Molecular Technique: First report from Pakistan

Objective: To evaluate bla SHV, bla TEM and bla OXA encoding clinical isolates in chronic tonsillitis using phenotypic and molecular techniques.Place and Duration:  The study was conducted in Department of Microbiology and Molecular Genetics, University of Punjab, Lahore from August 2016 to August 2017.Methodology: Sample processing, identification and characterization of isolates was done by using (CLSI, 2016) criteria. Antibiotic susceptibility testing by using disc diffusion assay and biofilm forming ability was analyzed by ring test and slime production test. Combination disc test was used for phenotypic detection of antibiotic resistance genes. Multiplex-PCR assay was used to check the presence of bla SHV, bla TEM and bla OXA genes. 16S rRNA sequencing and phylogenetic analysis was performed.Results: Here, variable resistance pattern was observed against applied antibiotics. 100 % resistance towards azotreonam and penicillin was observed. While 60-85 % resistances were observed against cephalosporins. Biofilm formation increased with the passage of time. 77% strains indicated positive combination disc test. Multiplex-PCR indicated 60% strains harbored tested genes. 40 % bla SHV genes, 30 % bla TEM genes and 60% bla OXA genes were observed among selected isolates. GenBank Accession number obtained for Klebsiella pneumoniae was KY810693 and for S. aureus was KY810692.Conclusion: In conclusion, K. pneumoniae and S. aureus came out to be common causative agents of tonsillitis in the current study. Resistance towards multiple classes of antibiotics and strong biofilms of these micro-organisms explain the chronicity and recurrent nature of the infection. bla OXA genes were frequent among genes tested

Size- and Shape-Dependent Antibacterial Studies of Silver Nanoparticles Synthesized by Wet Chemical Routes

Silver nanoparticles (AgNPs) of different shapes and sizes were prepared by solution-based chemical reduction routes. Silver nitrate was used as a precursor, tri-sodium citrate (TSC) and sodium borohydride as reducing agents, while polyvinylpyrrolidone (PVP) was used as a stabilizing agent. The morphology, size, and structural properties of obtained nanoparticles were characterized by scanning electron microscopy (SEM), UV-visible spectroscopy (UV-VIS), and X-ray diffraction (XRD) techniques. Spherical AgNPs, as depicted by SEM, were found to have diameters in the range of 15 to 90 nm while lengths of the edges of the triangular particles were about 150 nm. The characteristic surface plasmon resonance (SPR) peaks of different spherical silver colloids occurring in the wavelength range of 397 to 504 nm, whereas triangular particles showed two peaks, first at 392 nm and second at 789 nm as measured by UV-VIS. The XRD spectra of the prepared samples indicated the face-centered cubic crystalline structure of metallic AgNPs. The in vitro antibacterial properties of all synthesized AgNPs against two types of Gram-negative bacteria, Pseudomonas aeruginosa and Escherichia coli were examined by Kirby–Bauer disk diffusion susceptibility method. It was noticed that the smallest-sized spherical AgNPs demonstrated a better antibacterial activity against both bacterial strains as compared to the triangular and larger spherical shaped AgNPs

Bacillus cereus biofilm formation on central venous catheters of hospitalised cardiac patients

Formation of bacterial biofilms is a risk with many in situ medical devices. Biofilm-forming Bacillus species are associated with potentially life-threatening catheter-related blood stream infections in immunocompromised patients. Here, bacteria were isolated from biofilm-like structures within the lumen of central venous catheters (CVCs) from two patients admitted to cardiac hospital wards. Isolates belonged to the Bacillus cereus group, exhibited strong biofilm formation propensity, and mapped phylogenetically close to the B. cereus emetic cluster. Together, whole genome sequencing and quantitative PCR confirmed that the isolates constituted the same strain and possessed a range of genes important for and up-regulated during biofilm formation. Antimicrobial susceptibility testing demonstrated resistance to trimethoprim-sulphamethoxazole, clindamycin, penicillin and ampicillin. Inspection of the genome revealed several chromosomal beta-lactamase genes and a sulphonamide resistant variant of folP. This study clearly shows that B. cereus persisting in hospital ward environments may constitute a risk factor from repeated contamination of CVCs

Antibiotic Drug Resistance Pattern of Uropathogens in Pediatric Patients in Pakistani Population

The common prevalent diseases in the age of 0 to 6 are related to urinary tract infections. If not properly diagnosed, they will lead to urological and nephrological complications. Uropathogens are developing resistance against most drugs and are harder to treat. A study was done on the inpatients and outpatients of the two hospitals located in Lahore. A total of 39,750 samples that were both male and female were collected. Escherichia and Klebsiella were found in 234 samples based on biochemical characterization, growth on CLED agar, and white blood cell/pus cell (WBC) microscopy. In comparison to males, female samples had a higher number of uropathogens (1:1.29). From the samples of Shaikh Zayed Hospital (SZH), the ratio of Klebsiella to Escherichia (1:1.93) was reported, while this ratio was 1.84:1 from the Children Hospital (CH). The incidence of UTI was higher in the month of September. Randomly selected Escherichia and Klebsiella were verified via a 16S rRNA sequence. Antibiotic resistance profiling of isolated bacterial strains was done against 23 antibiotics. The most efficient antibiotics against Klebsiella and Escherichia were colistin sulphate (100% sensitivity against bacteria from CH; 99.3% against strains from SZH) and polymyxin B (100% sensitivity against strains from SZH; 98.8% against strains from CH). Sensitivity of the total tested strains against meropenem (74%, SZH; 70% CH), Fosfomycin (68%, SZH; 73% CH strains), amikacin (74% SZH; 55% CH), and nitrofurantoin (71% SZH;67% CH) was found, Amoxicillin, ampicillin, and cefuroxime showed 100 to ≥90% resistance and are the least effective