Prevalence of NDM‐1 carbapenemase in patients with diarrhoea in Pakistan and evaluation of two chromogenic culture media

Aims

To evaluate two chromogenic media, Brilliance CRE and chromID CARBA, with stool samples referred to the Public Health Laboratories Division of the National Institute of Health in Islamabad, and assess the prevalence of carbapenemase‐producing Enterobacteriaceae (CPE) in this population.

Methods and Results

One hundred and fifty‐two stool samples from patients with diarrhoea were referred to the Microbiology Department and were investigated for the presence of CPE using two chromogenic culture media, Brilliance CRE and chromID CARBA. Thirteen patients (8·6%) were found to be colonized with CPE and all produced NDM‐1 carbapenemase. Twelve of these patients (92%) were found to be colonized by culture on chromID CARBA compared with seven (54%) using Brilliance CRE.

Conclusions

If only coloured colonies were considered as presumptive CPE, the sensitivity, specificity and positive predictive value were 54, 23 and 6% for Brilliance CRE and 85, 85 and 36% for chromID CARBA, respectively.

Significance and Impact of the Study

We conclude that Enterobacteriaceae that produce NDM‐1 carbapenemase can be found in patients from all major provinces of Pakistan and that chromID CARBA was the most effective of the two chromogenic media in this setting.     

An efficient and rapid regeneration via multiple shoot induction from mature seed derived embryogenic and organogenic callus of Indian maize (Zea mays L.)

An efficient method for in vitro micro propagation and genetic transformation of plants are crucial for both basic and applied research. Maize is one of the most important cereal crops around the world. Regeneration from immature embryo is hampered due to its unavailability round the year. On the contrary mature embryo especially tropical maize is recalcitrant toward tissue culture. Here we report a highly efficient regeneration (90%) system for maize by using 2 different approaches i.e., embryogenic and organogenic callus cultures. Seeds were germinated on MS medium supplemented with 5 mg/l 2,4-D and 3 mg/l BAP. Nodal regions of 2 wks old seedlings were longitudinally split upon isolation and subsequently placed on callus initiation medium. The maximum frequency of embryogenic callus formation (90%) was obtained on MS medium supplemented with 2 mg/l 2,4-D and 1 mg/l BAP in the dark conditions. The compact granular organogenic callus formation (85% frequency) was obtained on MS medium supplemented with 2.5 mg/l 2,4-D and 1.5 mg/l BAP at light conditions. MS medium supplemented with 2 mg/l BAP, 1 mg/l Kinetin and 0.5 mg/l NAA promoted the highest frequency of shoot induction. The highest frequency of root formation was observed when shoots were grown on MS medium. The regenerated plants were successfully hardened in earthen pots after adequate acclimatization. The important advantage of this improved method is shortening of regeneration time by providing an efficient and rapid regeneration tool for obtaining more stable transformants from mature seeds of Indian tropical maize cultivar (HQPM-1).     

The Conserved Intronic Cleavage and Polyadenylation Site of CstF-77 Gene Imparts Control of 3′ End Processing Activity through Feedback Autoregulation and by U1 snRNP

The human gene encoding the cleavage/polyadenylation (C/P) factor CstF-77 contains 21 exons. However, intron 3 (In3) accounts for nearly half of the gene region, and contains a C/P site (pA) with medium strength, leading to short mRNA isoforms with no apparent protein products. This intron contains a weak 5′ splice site (5′SS), opposite to the general trend for large introns in the human genome. Importantly, the intron size and strengths of 5′SS and pA are all highly conserved across vertebrates, and perturbation of these parameters drastically alters intronic C/P. We found that the usage of In3 pA is responsive to the expression level of CstF-77 as well as several other C/P factors, indicating it attenuates the expression of CstF-77 via a negative feedback mechanism. Significantly, intronic C/P of CstF-77 pre-mRNA correlates with global 3′UTR length across cells and tissues. In addition, inhibition of U1 snRNP also leads to regulation of the usage of In3 pA, suggesting that the C/P activity in the cell can be cross-regulated by splicing, leading to coordination between these two processes. Importantly, perturbation of CstF-77 expression leads to widespread alternative cleavage and polyadenylation (APA) and disturbance of cell proliferation and differentiation. Thus, the conserved intronic pA of the CstF-77 gene may function as a sensor for cellular C/P and splicing activities, controlling the homeostasis of CstF-77 and C/P activity and impacting cell proliferation and differentiation.     

Mapping of durable stripe rust resistance in a durum wheat cultivar Wollaroi

Wollaroi, an Australian durum wheat cultivar, produced a low stripe rust response and the alternative parent Bansi was highly susceptible. The Wollaroi/Bansi recombinant inbred line (RIL) population was phenotyped across three consecutive crop seasons. A genetic map of the Wollaroi/Bansi RIL population comprising 799 markers (diversity arrays technology and simple sequence repeat markers) was used to determine the genomic location of stripe rust resistance genes carried by the cultivar Wollaroi. Composite interval mapping detected three consistent quantitative trait loci (QTL) in chromosomes 2A, 3B and 5B. These QTL were named QYr.sun-2A, QYr.sun-3B and QYr.sun-5B. Another QTL, QYr.sun-1B, was detected only in the 2009 crop season. QTL in chromosomes 1B, 2A, 3B and 5B explained on average 6, 9.3, 26.7 and 8.7 %, respectively, of the variation in stripe rust response. All QTL were contributed by Wollaroi. RILs carrying these QTL singly produced intermediate stripe rust severities ranging from 46.2 to 55.7 %, whereas RILs with all four QTL produced the lowest disease severity (34.3 %). The consistently low stripe rust response of Wollaroi for 20 years demonstrated the durability of the resistance loci involved. The QTL combination detected in this study is being transferred to common wheat.     

Cervical Cancer Screening among University Students in South Africa: A Theory Based Study

Introduction

Cervical cancer is a serious public health problem in South Africa. Even though the screening is free in health facilities in South Africa, the Pap smear uptake is very low. The objective of the study is to investigate the knowledge and beliefs of female university students in South Africa.

Methods

A cross sectional study was conducted among university women in South Africa to elicit information about knowledge and beliefs, and screening history.

Results

A total of 440 students completed the questionnaire. The average age of the participants was 20.39 years (SD  = 1.71 years). Regarding cervical cancer, 55.2% (n = 243) had ever heard about it. Results indicated that only 15% (22/147) of the students who had ever had sex and had heard about cervical cancer had taken a Pap test. Pearson correlation analysis showed that cervical cancer knowledge had a significantly negative relationship with barriers to cervical cancer screening. Susceptibility and seriousness score were significantly moderately correlated with benefit and motivation score as well as barrier score. Self-efficacy score also had a moderate correlation with benefit and motivation score. Students who had had a Pap test showed a significantly lower score in barriers to being screened compared to students who had not had a Pap test.

Conclusion

This study showed that educated women in South Africa lack complete information on cervical cancer. Students who had had a Pap test had significantly lower barriers to cervical cancer screening than those students who had not had a Pap test.     

Interaction Between Essential Elements Selenium and Zinc with Cadmium and Mercury in Samples from Hypertensive Patients

The abnormal metabolism of metal ions plays an important role in health and disease conditions; hence, the studies about them have received much interest. The objective of this study was to evaluate the association between trace and toxic elements zinc (Zn), cadmium (Cd), selenium (Se), and mercury (Hg) in biological samples (scalp hair, blood, and urine) of hypertensive patients (n?=?257), residents of Hyderabad, Pakistan. For comparison purpose, the biological samples of age-matched healthy controls were selected as referents. The concentrations of trace and toxic elements were measured by atomic absorption spectrophotometer prior to microwave-assisted acid digestion. The validity and accuracy of the methodology was checked using certified reference materials and by the conventional wet acid digestion method. The recovery of all studied elements was found in the range of 96.4–99.1 % in certified reference materials. The results of this study showed that the mean values of Cd and Hg were significantly higher in scalp hair, blood, and urine samples of hypertensive patients than in referents (P?<?0.001), whilst the concentrations of Zn and Se were lower in the scalp hair and blood, but higher in the urine samples of hypertensive patients. The deficiency of Zn and Se and the high exposure of toxic metals may be synergistic with risk factors associated with hypertension.     

Nuclear factor-kappaB p65 inhibits mitogen-activated protein kinase signaling pathway in radioresistant breast cancer cells

The molecular mechanism by which tumor cells increase their resistance to therapeutic radiation remains to be elucidated. We have previously reported that activation of nuclear factor-kappaB (NF-kappaB) is causally associated with the enhanced cell survival of MCF+FIR cells derived from breast cancer MCF-7 cells after chronic exposure to fractionated ionizing radiation. The aim of the present study was to reveal the context of NF-kappaB pathways in the adaptive radioresistance. Using cell lines isolated from MCF+FIR populations, we found that the elevated NF-kappaB activity was correlated with enhanced clonogenic survival, and increased NF-kappaB subunit p65 levels were associated with a decrease in phosphorylation of mitogen-activated protein kinase/extracellular signal-regulated kinase (ERK) kinase (MEK)/ERK in all radioresistant MCF+FIR cell lines. Further irradiation with 30 fractions of radiation also inhibited MEK/ERK phosphorylation in paired cell lines of MCF+FIR and parental MCF-7 cells. Activation of ataxia-telangiectasia mutated (ATM) protein, a sensor to radiation-induced DNA damage, was elevated with increased interaction with NF-kappaB subunits p65 and p50. The interaction between p65 and MEK was also enhanced in the presence of activated ATM. In contrast, both interaction and nuclear translocation of p65/ERK were reduced. Inhibition of NF-kappaB by overexpression of mutant IkappaB increased ERK phosphorylation. In addition, MEK/ERK inhibitor (PD98059) reduced the interaction between p65 and ERK. Taken together, these results suggest that NF-kappaB inhibits ERK activation to enhance cell survival during the development of tumor adaptive radioresistance.     

Targeted cross-linking of the human beta-globin gene in living cells mediated by a triple helix forming oligonucleotide

Triple helix forming oligonucleotides (TFOs) may have utility as gene targeting reagents for "in situ" gene therapy of genetic disorders. Triplex formation is challenged by negative charge repulsion between third strand and duplex phosphates, and destabilizing positive charge repulsion between adjacent protonated cytosines within pyrimidine motif third strands. Here we describe the synthesis of TFOs designed to target a site in the human beta-globin gene, which is the locus for mutations that underlie the beta-globinopathies, including sickle cell anemia. The target is an uninterrupted polypurine:polypyrimidine sequence, containing four adjacent cytosines, next to a psoralen cross-link site. Pyrimidine motif TFOs that contained four adjacent cytosines or 5-methylcytosines did not form stable triplexes at physiological pH, despite the introduction of otherwise stabilizing base and sugar analogues. We synthesized a series of pso-TFOs containing 2'-O-methyl (OMe) and 2'-O-aminoethoxy substitutions (AE), as well as 8-oxo-adenine (A8) and 2'-O-methylpseudoisocytidine (P) as neutral cytosine replacements. Thermal stability measurements indicated that TFOs with A8 did not meet criteria established in previous work. However, TFOs with P did form triplexes with appropriate T(m) and k(ON) values. A pso-TFO with AE and P residues was sufficiently active to permit the determination of targeting in living cells by direct measurement of cross-link formation at the target site. Our results validate the modification format described in our previous studies and indicate that P substitutions are an effective solution to the problem of targeting genomic sequences containing adjacent cytosines.