Frequency of conjugative transfer of plasmid-encoded ISEcp1 - blaCTX-M-15 and aac(6')-lb-cr genes in Enterobacteriaceae at a tertiary care center in Lebanon - role of transferases

<p>Abstract</p> <p>Background</p> <p>The frequency of transfer of genes encoding resistance to antimicrobial agents was determined by conjugation in ESBL-producing and/or fluoroquinolone or aminoglycoside resistant Enterobacteriaceae clinical isolates at a tertiary care center in Lebanon. In addition, the role of <it>tra </it>genes encoding transferases in mediating conjugation was assessed.</p> <p>Methods</p> <p>Conjugation experiments were done on 53 ESBL-producing and/or fluoroquinolone resistant <it>E. coli </it>and <it>K. pneumoniae </it>and ESBL-producing <it>S. sonnei </it>isolates. Antimicrobial susceptibility testing on parent and transconjugant isolates, and PCR amplifications on plasmid extracts of the resistance-encoding genes: <it>bla</it>_CTX-M-15with the <it>ISEcp1 </it>insertion sequence, the <it>aac(6')-lb-cr </it>and <it>qnr</it>S genes, as well as <it>tra </it>encoding transferases genes were done. Random amplified polymorphic DNA (RAPD) analysis was performed to demonstrate whether conjugative isolates are clonal and whether they are linked epidemiologically to a particular source.</p> <p>Results</p> <p>Antimicrobial susceptibility testing on transconjugants revealed that 26 out of 53 (49%) ESBL-producing <it>Enterobacteriaceae </it>were able to transfer antimicrobial resistance to the recipients. Transfer of high-level resistance to the transconjugants encoded by the <it>bla</it>_CTX-M-15gene downstream the <it>ISEcp1 </it>insertion sequence against 3rd generation cephalosporins, and of low-level resistance against ciprofloxacin, and variable levels of resistance against aminoglycosides encoded by <it>aac(6')-lb-cr </it>gene, were observed in transconjugants. <it>tra </it>encoding transferase genes were detected exclusively in conjugative isolates.</p> <p>Conclusion</p> <p>In conclusion, the frequency of transfer of antimicrobial resistance in non clonal <it>Enterobacteriaceae </it>at the tertiary care center by conjugation was 49%. Conjugation occurred in isolates expressing the <it>tra </it>encoding transferase genes. Multiple conjugative strains harboring the plasmid encoded antimicrobial resistant genes were circulating in the medical center. Molecular epidemiology analysis showed that conjugative isolates are neither clonal nor linked to a particular site and transfer of antimicrobial resistance is by horizontal transfer of plasmids.</p

Correlation between Group B Streptococcal Genotypes, Their Antimicrobial Resistance Profiles, and Virulence Genes among Pregnant Women in Lebanon

The antimicrobial susceptibility profiles of 76 Streptococcus agalactiae (Group B Streptococci [GBS]) isolates from vaginal specimens of pregnant women near term were correlated to their genotypes generated by Random Amplified Polymorphic DNA analysis and their virulence factors encoding genes cylE, lmb, scpB, rib, and bca by PCR. Based on the distribution of the susceptibility patterns, six profiles were generated. RAPD analysis detected 7 clusters of genotypes. The cylE gene was present in 99% of the isolates, the lmb in 96%, scpB in 94.7%, rib in 33%, and bca in 56.5% of isolates. The isolates demonstrated a significant correlation between antimicrobial resistance and genotype clusters denoting the distribution of particular clones with different antimicrobial resistance profiles, entailing the practice of caution in therapeutic options. All virulence factors encoding genes were detected in all seven genotypic clusters with rib and bca not coexisting in the same genome

Genotypes and serotype distribution of macrolide resistant invasive and non- invasive Streptococcus pneumoniae isolates from Lebanon

<p>Abstract</p> <p>Background</p> <p>This study determined macrolide resistance genotypes in clinical isolates of <it>Streptococcus pneumoniae </it>from multiple medical centers in Lebanon and assessed the serotype distribution in relation to these mechanism(s) of resistance and the source of isolate recovery.</p> <p>Methods</p> <p>Forty four macrolide resistant and 21 macrolide susceptible <it>S. pneumoniae </it>clinical isolates were tested for antimicrobial susceptibility according to CLSI guidelines (2008) and underwent molecular characterization. Serotyping of these isolates was performed by Multiplex PCR-based serotype deduction using CDC protocols. PCR amplification of macrolide resistant <it>erm </it>(encoding methylase) and <it>mef </it>(encoding macrolide efflux pump protein) genes was carried out.</p> <p>Results</p> <p>Among 44 isolates resistant to erythromycin, 35 were resistant to penicillin and 18 to ceftriaxone. Examination of 44 macrolide resistant isolates by PCR showed that 16 isolates harbored the <it>erm</it>(B) gene, 8 isolates harbored the <it>mef </it>gene, and 14 isolates harbored both the <it>erm</it>(B) and <it>mef </it>genes. There was no amplification by PCR of the <it>erm</it>(B) or <it>mef </it>genes in 6 isolates. Seven different capsular serotypes 2, 9V/9A,12F, 14,19A, 19F, and 23, were detected by multiplex PCR serotype deduction in 35 of 44 macrolide resistant isolates, with 19F being the most prevalent serotype. With the exception of serotype 2, all serotypes were invasive. Isolates belonging to the invasive serotypes 14 and 19F harbored both <it>erm</it>(B) and <it>mef </it>genes. Nine of the 44 macrolide resistant isolates were non-serotypable by our protocols.</p> <p>Conclusion</p> <p>Macrolide resistance in <it>S. pneumoniae </it>in Lebanon is mainly through target site modification but is also mediated through efflux pumps, with serotype 19F having dual resistance and being the most prevalent and invasive.</p

Treated municipal wastewater reuse for eggplant irrigation

In the Bekaa Valley of Lebanon, the availability of freshwater is limited and therefore farmers must start using alternative water sources such as treated wastewater for irrigating crops. The present study is of great importance, since it provides evidence of some on-farm options that farmers can adopt when irrigating with treated effluent, in order to minimize the health risks. Eggplant was grown under two water quality regimes (Freshwater (FW) and treated wastewater (TW)) and two agronomic practices (no mulch (N_Mu) and use of plastic mulch (Mu)). Treatments were arranged in a split plot design with four replicates. Water quality regime was the main plot factor, while agronomic practices were the subplot factors. Water quality, soil, the marketable yield and other parameters were measured. Fruit samples were evaluated for bacterial contamination. The drip line performance was monitored. The study results revealed that the treatment with treated effluent gave more fruits.m-2, yield and mean fruit weight than the treatment using fresh water irrigation, with an increase of 3.98%, 10.74% and 5.63%, respectively. In addition, the use of mulch (Mu) resulted in an increase in yield (24.23%) and number of fruits (14.11%). Concerning the emitters' performance and sensitivity to clogging, discharge reduction rate (Rd = 6.75%) for drippers delivering TW was lower than the admissible value of 20% discharge variation, indicating that the quality of water has little effect on emitter performance. Concerning bacterial contamination of fruits, irrigation with treated wastewater showed no contamination in terms of fecal streptococci, salmonella or E. coli. However, the fruits were contaminated with fecal coliforms that were present at a concentration less than 200 CFU.100 g-1. Following the World Health Organization Guidelines, pathogens could be reduced through post treatment health-protection control measures such as drip irrigation, product washing, disinfection and produce peelin

Contribution of the immune system and astrocytes to spinal muscular atrophy pathology

Field of study: Microbiology and immunology.Dr. Christian L. Lorson, Dissertation Supervisor.Includes vita."May 2018."[ACCESS RESTRICTED TO THE UNIVERSITY OF MISSOURI AT AUTHOR'S REQUEST.] Spinal muscular atrophy (SMA) is an autosomal recessive neuromuscular disease and the leading genetic cause of infant mortality. SMA is caused by afunctional loss of the survival motor neuron-1 (SMN1) gene and the subsequent deficiency of the ubiquitously expressed survival motor neuron (SMN) protein. SMA presents by motor neuron loss and muscle atrophy, and historically was considered an autonomous disease of the a-lower motor neuron (LMN). In this work we investigated effects of low levels of SMN outside the LMNs. Specifically, we looked in the spleen to determine the impact of SMN deficiency on the spleen development and the integrity of the splenic immune cells. Additionally, we analyzed astrocytes to determine if they exhibit functional impairments that could compromise their role in supporting the survival and function of LMNs. First, we reported spleen hypoplasia in multiple SMA mouse models with alteration of the splenic architecture due to a severe reduction in the red pulp zone and relative conservation of the white pulp area. We found alterations in the relative abundance of splenic mediators of the immune response, where the resident macrophage subset was depleted starting at early pre-symptomatic time (post-natal day 2), whereas the B- and T-lymphocytes, and CD11b+ macrophages had higher frequency at the late symptomatic age (post-natal day 12). Secondly, we showed that primary astrocytes derived from the spinal cord of a SMA mouse model had compromised efficiency in their glutamate uptake capacity. Moreover, SMA spinal cords had altered expression in the level of flottilin-1, a lipid raft protein necessary for cell membrane integrity and for the function of receptors and transporters. Furthermore, we showed a higher expression of the NR2B, a subunit of the glutamate receptor NMDAR that signals mainly toward apoptosis. In summary, this work characterized new pathologies in two non-neuronal tissues in the CNS and in the periphery, demonstrated that the spleen and the immune system are likely contributing to the overall clinical pathology of SMA, and found altered mechanisms in astrocytes function that might explain their effect on LMNs in SMA.Includes bibliographical references (pages 98-116)

Application of sewage sludge for cereal production in a Mediterranean environment (Lebanon)

Purpose Management of sewage sludge generated from wastewater treatment plants is a big challenge for its potential reusage in agriculture. Most of the Lebanese local sludge is discarded in the water or in landfills (most wastewater plants are partially functional). The objective of this research was to assess the effect of the application of different sewage sludge rates on the wheat production as an alternative of chemical fertilizer. Method Field trials were conducted, for one-year study, in IAAT village in the Bekaa valley-Lebanon. The considered treatments of 4, 8 and 16 kg.m-2 rates were compared to a control treatment. Physicochemical and microbiological analysis were performed on sludge and soil samples (pre cultivation and post-harvest). The harvested wheat was also analyzed for several parameters as mineral content. Results Results presented significant differences between control and treatments. pH values decreased with biosolids additions; organic matter rose in the amended soils, macronutrients levels increased. Heavy metals outcome increased significantly after amendment, microbiological analysis didn’t show any contamination by Salmonella, E.Coli, Staphylococcus Aureus and Helminth eggs. As for wheat plants' evaluation, fiber and protein contents presented an increase similar to nitrogen and phosphorus. Conclusion These results are a key component that identifies the role of biosolids as pH regulator and soil conditioner which improves the physicochemical properties of soil without any risk of microbiological contamination. These results are promising and they encourage the use of biosolids as agriculture amendment

SMN deficiency negatively impacts red pulp macrophages and spleen development in mouse models of spinal muscular atrophy

Spinal muscular atrophy (SMA) is a progressive neurodegenerative disease that is the leading genetic cause of infantile death. It is caused by a severe deficiency of the ubiquitously expressed Survival Motor Neuron (SMN) protein. SMA is characterized by α-lower motor neuron loss and muscle atrophy, however, there is a growing list of tissues impacted by a SMN deficiency beyond motor neurons. The non-neuronal defects are observed in the most severe Type I SMA patients and most of the widely used SMA mouse models, however, as effective therapeutics are developed, it is unclear whether additional symptoms will be uncovered in longer lived patients. Recently, the immune system and inflammation has been identified as a contributor to neurodegenerative diseases such as ALS. To determine whether the immune system is comprised in SMA, we analyzed the spleen and immunological components in SMA mice. In this report, we identify: a significant reduction in spleen size in multiple SMA mouse models and a pathological reduction in red pulp and extramedullary hematopoiesis. Additionally, red pulp macrophages, a discrete subset of yolk sac-derived macrophages, were found to be altered in SMA spleens even in pre-symptomatic post-natal day 2 animals. These cells, which are involved in iron metabolism and the phagocytosis of erythrocytes and blood-borne pathogens are significantly reduced prior to the development of the neurodegenerative hallmarks of SMA, implying a differential role of SMN in myeloid cell ontogeny. Collectively, these results demonstrate that SMN deficiency impacts spleen development and suggests a potential role for immunological development in SMA

In trans T cell tolerance exacerbates experimental allergic encephalomyelitis by interfering with protective antibody responses

Despite being one of the most biodiverse regions in the world, not much is known concerning the ecology of the otters on Borneo. We conducted a study to document the activity patterns of the smooth-coated otter, Lutrogale perspicillata, in increasingly disturbed and fragmented habitats in the Lower Kinabatangan Wildlife Sanctuary (LKWS), located in the Malaysian state of Sabah, northern Borneo. The aim was to gather ecological information for establishing baseline data and to understand better the otter behavior in this region of Sabah. We deployed camera traps at active otter holts, grooming and sprainting sites for 15 non-consecutive months and utilized the photographs to model the activity patterns of the otterusing kernel’s density estimate modeling. Results showed that L. perspicillatain the LKWS was mainly crepuscular, with otter activity mainly occurring during early morning (0600 h) and late afternoon (1600 h -1800 h). Grooming activity peaked at 0600 h while sprainting activity peaked at both 0800h and 1700 h. We suggest that activity patterns of L. perspicillatamay be influenced by prey availability, human disturbance and environmental temperature