MicroRNA characterize genetic diversity and drug resistance in pediatric acute lymphoblastic leukemia

Background

MicroRNA regulate the activity of protein-coding genes including those involved in hematopoietic cancers. The aim of the current study was to explore which microRNA are unique for seven different subtypes of pediatric acute lymphoblastic leukemia.

Design and Methods

Expression levels of 397 microRNA (including novel microRNA) were measured by quantitative real-time polymerase chain reaction in 81 cases of pediatric leukemia and 17 normal hematopoietic control cases.

Results

All major subtypes of acute lymphoblastic leukemia, i.e. T-cell, MLL-rearranged, TEL-AML1-positive, E2A-PBX1-positive and hyperdiploid acute lymphoblastic leukemia, with the exception of BCR-ABL-positive and ‘B-other’ acute lymphoblastic leukemias (defined as precursor B-cell acute lymphoblastic leukemia not carrying the foregoing cytogenetic aberrations), were found to have unique microRNA-signatures that differed from each other and from those of healthy hematopoietic cells. Strikingly, the microRNA signature of TEL-AML1-positive and hyperdiploid cases partly overlapped, which may suggest a common underlying biology. Moreover, aberrant down-regulation of let-7b (~70-fold) in MLL-rearranged acute lymphoblastic leukemia was linked to up-regulation of oncoprotein c-Myc (P_FDR<0.0001). Resistance to vincristine and daunorubicin was characterized by an approximately 20-fold up-regulation of miR-125b, miR-99a and miR-100 (P_FDR≤0.002). No discriminative microRNA were found for prednisolone response and only one microRNA was linked to resistance to L-asparaginase. A combined expression profile based on 14 microRNA that were individually associated with prognosis, was highly predictive of clinical outcome in pediatric acute lymphoblastic leukemia (5-year disease-free survival of 89.4%±7% versus 60.8±12%, P=0.001).

Conclusions

Genetic subtypes and drug-resistant leukemic cells display characteristic microRNA signatures in pediatric acute lymphoblastic leukemia. Functional studies of discriminative and prognostically important microRNA may provide new insights into the biology of pediatric acute lymphoblastic leukemia.     

VH gene mutation status and cellular drug resistance in chronic lymphocytic leukaemia

OBJECTIVE: B-cell chronic lymphocytic leukaemia (B-CLL) can be divided into two clinical entities based on the immunoglobulin variable heavy chain (VH) gene mutation status, as cases with unmutated VH genes display a more aggressive disease with shorter survival time than mutated cases. The aim of this study was to investigate whether differences in cellular drug resistance could give an explanation for these divergent clinical courses. METHODS: The VH gene mutation status was analysed in patients with previously untreated B-CLL using VH gene family-specific PCR amplification and nucleotide sequencing. In vitro sensitivity to cytarabine, fludarabine, cladribine, doxorubicin, idarubicin, vincristine, cyclophosphamide, melphalan and prednisolone was assessed using the non-clonogenic in vitro assay, fluorometric microculture cytotoxicity assay. RESULTS: The VH genes and in vitro drug resistance were successfully analysed in 46 cases, revealing that 25 (54%) cases showed unmutated and 21 (46%) cases mutated VH genes. Interestingly, the unmutated group generally tended to be more chemosensitive than the mutated group with significant differences for cytarabine and prednisolone (P < or = 0.01). CONCLUSION: The propensity of inferior drug response in mutated B-CLL may reflect a more differentiated disease than in unmutated B-CLL. We conclude that the difference in prognosis between B-CLL cases with unmutated and mutated VH genes could not be explained by difference in cellular drug resistance.     

粪肠球菌gyrA基因突变与氟喹诺酮类耐药性关系的研究

目的探讨粪肠球菌DNA促旋酶A亚单位（gyrA）基因突变与氟嚎诺酮类耐药的关系。方法琼脂稀释法测定粪肠球菌对各种氟喹诺酮类药物的耐药情况;聚合酶链反应（PCR）结合单链构象多态性（SSCP）分析检测gyrA中氟哇诺酮类耐药决定区（QRDR）突变情况。结果粪肠球菌对诺氟沙星耐药率为78．1％（50／64）,加替沙星为54．7％（35／64）。41株耐药菌的第87位氨基酸编码碱基发生突变（GAA→GGA）。结论加替沙星的耐药率较低,优于其他氟哇诺酮类药物;gyrA基因突变是粪肠球菌对氟喹诺酮类耐药的重要原因。     

AcrB drug-binding pocket substitution confers clinically relevant resistance and altered substrate specificity

The incidence of multidrug-resistant bacterial infections is increasing globally and the need to understand the underlying mechanisms is paramount to discover new therapeutics. The efflux pumps of Gram-negative bacteria have a broad substrate range and transport antibiotics out of the bacterium, conferring intrinsic multidrug resistance (MDR). The genomes of pre- and posttherapy MDR clinical isolates of Salmonella Typhimurium from a patient that failed antibacterial therapy and died were sequenced. In the posttherapy isolate we identified a novel G288D substitution in AcrB, the resistance-nodulation division transporter in the AcrAB-TolC tripartite MDR efflux pump system. Computational structural analysis suggested that G288D in AcrB heavily affects the structure, dynamics, and hydration properties of the distal binding pocket altering specificity for antibacterial drugs. Consistent with this hypothesis, recreation of the mutation in standard Escherichia coli and Salmonella strains showed that G288D AcrB altered substrate specificity, conferring decreased susceptibility to the fluoroquinolone antibiotic ciprofloxacin by increased efflux. At the same time, the substitution increased susceptibility to other drugs by decreased efflux. Information about drug transport is vital for the discovery of new antibacterials; the finding that one amino acid change can cause resistance to some drugs, while conferring increased susceptibility to others, could provide a basis for new drug development and treatment strategies.The incidence of multidrug-resistant (MDR, also used herein for “multidrug resistance”) bacterial infections is increasing, and the 2013 World Economic Forum Global Risks report listed antibiotic-resistant bacteria as one of the greatest threats to human health (1). Resistance-nodulation division (RND) efflux pumps of Gram-negative bacteria confer intrinsic and acquired MDR in clinically relevant infections by exporting antibiotics out of the bacterial cell, allowing bacteria to survive at increased drug concentrations.To date the best-characterized efflux pump is AcrAB-TolC of Escherichia coli, composed of the inner membrane RND antiporter AcrB that functions in a tripartite assembly with a periplasmic adaptor protein, AcrA, and the outer membrane channel, TolC. The AcrB pump is an asymmetric homotrimer whose monomers undergo a functional rotation through three states: access, binding, and extrusion (labeled A, B, and E, respectively) to pump substrates from the periplasm, or outer leaflet of the inner membrane, to outside of the cell (2, 3). Drug binding within AcrB is complex. The distal binding pocket has a phenylalanine-rich region that binds low-molecular-weight drugs and a proximal binding pocket and vestibule bind larger compounds such as erythromycin (4, 5).To date, MDR in clinically relevant infections mediated by RND efflux pumps, including AcrB, has been documented to be due to overexpression of the efflux pump and concomitant increased efflux of antibiotics from the bacterial cell (6). Overproduction of efflux pumps can occur via four mechanisms: (i) mutation of the local repressor gene (7, 8), (ii) mutation in a global regulatory gene (9, 10), (iii) mutation of the promoter region of the efflux pump gene (11), or (iv) insertion elements upstream of the transporter gene (12, 13).This study focused on elucidating the mechanism of resistance in a unique set of clinical isolates collected over the course of a complex Salmonella infection (Fig. S1) (14, 15). The isolates were obtained from a 52-y-old male patient admitted for repair of a leaking abdominal aortic aneurysm graft. Salmonella Typhimurium (L3) was isolated before ciprofloxacin treatment and was susceptible to ampicillin, sulphonamide, trimethoprim, cefuroxime, chloramphenicol, gentamicin, and ciprofloxacin. Over the course of the infection the patient received i.v. ciprofloxacin, oral ciprofloxacin, i.v. ceftazidime, and i.v. aztreonam. Isolates were taken throughout infection, and during treatment MDR strains were isolated. L18, the last MDR strain to be isolated, was from wound drainage fluid when the patient had received no antibiotics for 2 wks. The patient died soon afterward with the infection unresolved. Compared with the pretherapy isolate L3, the MDR posttherapy isolate, L18, was less susceptible to numerous agents, including ciprofloxacin and β-lactams, and accumulated less ciprofloxacin and Hoechst dye (16). This set of isolates has provided a unique opportunity to investigate in vivo evolution of MDR in response to clinically validated courses of antimicrobial treatment.Here, we report that whole genome sequencing revealed a new mechanism of clinically significant MDR selected during therapy: substitution in a transporter protein, which altered the specificity of the efflux pump for antibacterial drugs. The structural impact of the substitution was investigated by molecular dynamics (MD) simulations.     

Drug resistance pattern and mutation pattern in pediatric tuberculosis: Study from north India

BackgroundThe emergence of multidrug-resistant MDR-TB and extensively drug-resistant XDR-TB are serious threats to global TB control. Molecular tests like GenoType MTBDRplus has revolutionized MDR-TB diagnosis by rapid detection of resistance, leading to early and appropriate management of DR-TB. Information about common mutations imparting resistance to RIF and INH, helps in understanding the disease epidemiology in various regions. The study was conducted to determine the genetic mutation in drug resistant tuberculosis in children less than 12 years with pulmonary or extrapulmonary tuberculosis.Materials/methodsRetrospective analysis was done over a period of 54 months from January 2015 to June 2019 to study the resistance pattern and mutations present in DR-TB in children less than 12 years with suspected pulmonary or extrapulmonary tuberculosis using Hain's GenoType MTBDRplus VER 2.0.ResultsOver a period of 54 months, samples from 3461 patients with suspected TB were received for MGIT culture, out of which, 347 were positive for Mycobacterium tuberculosis. 250 of these 347 isolated were tested for drug resistance by Hain's GenoType MTBDRplus VER 2.0.61.1% were sensitive to isoniazid and rifampicin while 15.2% were DR-TB (38 out of 250). Out of these 38, 22 were MDR TB, 13 were isoniazid monoresistant (34.2%) and 3 were rifampicin monoresistant. The most common genotypic resistance for rifampicin was absence of rpoB WT8 band and presence of rpoB MUT 3 band (88%). 84.6% of the INH monoresistant isolates showed high level isoniazid resistant. All these isolates showed presence of katG MUT 1 band. On comparing Hain's GenoType MTBDRplus VER 2.0 with Xpert MTB/Rif Assay, most common mutation for rifampicin resistance at S531L which can be detected by Xpert MTB/Rif Assay (probe E). However, two cases with rifampicin resistance had mutation in codon region 509–513 and 513–519 which could be missed by Xpert MTB/Rif Assay.ConclusionsWe cannot solely rely on Xpert MTB/Rif Assay for detection of drug resistance due to the risk of missing the isoniazid monoresistance. GenoType MTBDRplus has revolutionized MDR-TB diagnosis by substantially reducing turn around time and leading to early management of DR-TB cases.     

Dried whole-plant Artemisia annua slows evolution of malaria drug resistance and overcomes resistance to artemisinin

Pharmaceutical monotherapies against human malaria have proven effective, although ephemeral, owing to the inevitable evolution of resistant parasites. Resistance to two or more drugs delivered in combination will evolve more slowly; hence combination therapies have become the preferred norm in the fight against malaria. At the forefront of these efforts has been the promotion of Artemisinin Combination Therapy, but despite these efforts, resistance to artemisinin has begun to emerge. In 2012, we demonstrated the efficacy of the whole plant (WP)—not a tea, not an infusion—as a malaria therapy and found it to be more effective than a comparable dose of pure artemisinin in a rodent malaria model. Here we show that WP overcomes existing resistance to pure artemisinin in the rodent malaria Plasmodium yoelii. Moreover, in a long-term artificial selection for resistance in Plasmodium chabaudi, we tested resilience of WP against drug resistance in comparison with pure artemisinin (AN). Stable resistance to WP was achieved three times more slowly than stable resistance to AN. WP treatment proved even more resilient than the double dose of AN. The resilience of WP may be attributable to the evolutionary refinement of the plant’s secondary metabolic products into a redundant, multicomponent defense system. Efficacy and resilience of WP treatment against rodent malaria provides compelling reasons to further explore the role of nonpharmaceutical forms of AN to treat human malaria.The fight against malaria predates the discovery of its causative agent, and for centuries malaria-associated fever was treated using herbal remedies. In the West, quinine (Cinchona bark extract) was the only affordable treatment against malaria until Paul Ehrlich’s magic bullet concept was adopted and thousands of synthetic compounds were tested against malaria parasites. Very few of these compounds were effective and/or safe for human use, but in the 1930s chloroquine rose to ascendancy as a miracle cure for malaria (1). In the late 1950s, chloroquine was the main weapon used by the World Health Organization (WHO) in its Global Malaria Eradication Program (GMEP). Sadly, development of drug-resistant parasites and concomitant failure of chloroquine as the drug of choice led to the eventual demise of GMEP by the close of the 1960s. Following chloroquine’s failure, various antimalarial compounds were serially deployed, and each in its turn failed as parasites evolved resistance, thus leaving millions of malaria patients without affordable treatment.In the 1970s, artemisinin was discovered as a pure drug extracted from the plant Artemisia annua. In wide-scale clinical trials, pure artemisinin showed poor pharmacokinetic properties but nonetheless demonstrated potent antimalarial activity with a high safety profile (2). It was determined that artemisinin when modified to artesunate or artemether improved bioavailability and was more effective when used in combination with other antimalarial drugs, mainly mefloquine, which became known as Artemisinin Combination Therapy (ACT) (3). It was hoped that use of ACTs would minimize risk of drug resistance. However, in 2005 the earliest evidence of P. falciparum resistance to ACTs arose in Southeast Asia (4–8). The fight against malaria became critical once again when it became apparent that ACT might be following chloroquine’s path toward obsolescence with no affordable replacement in sight.We demonstrated the efficacy of the whole A. annua plant as a malaria therapy and found it to be more effective than a comparable dose of pure artemisinin in a rodent malaria model (9). WHO has cautioned against use of nonpharmaceutical sources of artemisinin because of the risk of delivering subtherapeutic doses that could exacerbate the resistance problem (10). This warning is valid given the low artemisinin content of juice extractions, teas, and infusion preparations of plant material used for most nonpharmaceutical plant-based therapies. However, the Whole Plant (WP) A. annua therapy that we have tested is not an extraction, a tea, or an infusion, but is based on oral consumption of the dried leaves of the whole plant. Based on our proof of principle in a rodent model, we postulate that with further development WP might provide a more abundant and affordable source of artemisinin-based therapy by eliminating the need for artemisinin extraction during manufacture.WP may be more effective than monotherapeutic artemisinin because WP may constitute a naturally occurring combination therapy that augments artemisinin delivery and synergizes the drug’s activity. This plant Artemisinin Combination Therapy (pACT) is the result of evolutionary refinement of the plant’s secondary metabolic products into a resilient and multicomponent defense system. As was demonstrated for other combination therapies, we hypothesized that a WP-based pACT would (i) overcome existing resistance to monotherapeutic pure artemisinin and (ii) increase the longevity of this therapy by delaying the onset of parasite resistance among wild types. Here, we tested these two hypotheses in two mouse malaria models, artemisinin-resistant Plasmodium yoelii (strain: ART) and artemisinin-sensitive Plasmodium chabaudi (strain: ASS).     

18例抗病毒治疗失败儿童艾滋病患者耐药基因突变规律研究

目的对儿童艾滋病抗病毒治疗失败病例的耐药基因突变规律及特点进行研究。方法分析18例接受过高效抗逆转录病毒治疗并已经出现病毒学及免疫学失败儿童艾滋病患者的横断面临床及实验室资料,对其耐药突变结果进行分析。结果患者年龄11.6±2.4岁,治疗的时间为36±12个月,其中15例患者曾经接受过成人抗病毒药物治疗,患者的 CD4~ T 淋巴细胞为34±30个/μl(3～96个/μl),病毒载量为5.23±0.57 log10 copies/ml(4.27～6.53 log10 copies/ml)。耐药发生率为100%。对非核苷类逆转录酶抑制剂(NNRTIs)耐药突变结果分析显示:所有患者均对奈韦拉平(NVP)产生高度耐药,对依非韦伦(EFV)高度耐药的有16例。对核苷类逆转录酶抑制剂(NRTIs)耐药突变结果分析显示:对拉米夫定(LAM)高度耐药的有14例,对去羟肌苷(ddI)高度耐药的有11例,对齐多夫定(AZT)高度耐药的有14例,对司他夫定(d4T)高度耐药的有16例。重要突变位点包括 Y181C(9例)、K103N(7例)、G190A(8例)、TAMs(17例)、M184V(10例)、K65R(5例)、Q151M(2例)。结论该组治疗失败儿童患者对正在使用的 NRTIs 及 NNRTIs 均已产生高度耐药,须考虑更换新的抗病毒治疗方案。     

Baculovirus resistance in codling moth is virus isolate-dependent and the consequence of a mutation in viral gene pe38

The baculovirus Cydia pomonella granulovirus (CpGV) is widely applied as a biocontrol agent of codling moth. After field resistance of codling moth populations had been observed against the commercially used Mexican (M) isolate of CpGV, infection experiments of larvae of the resistant codling moth strain CpRR1 showed that several other naturally occurring CpGV isolates (I12, S, E2, and I07) from different geographic origins are still infectious to resistant CpRR1. Whole-genome sequencing and phylogenetic analyses of these geographic CpGV variants revealed that their genomes share only a single common difference from that of CpGV-M, which is a mutation coding for a repeat of 24 nucleotides within the gene pe38; this mutation results in an additional repeat of eight amino acids that appears to be inserted to PE38 of CpGV-M only. Deletion of pe38 from CpGV-M totally abolished virus infection in codling moth cells and larvae, demonstrating that it is an essential gene. When the CpGV-M deletion mutant was repaired with pe38 from isolate CpGV-S, which originated from the commercial product Virosoft and is infectious for the resistant codling moth strain CpRR1, the repaired CpGV-M mutant was found to be fully infectious for CpRR1. Repair using pe38 from CpGV-M restored infectivity for the virus in sensitive codling moth strains, but not in CpRR1. Therefore, we conclude that CpGV resistance of codling moth is directed to CpGV-M but not to other virus isolates. The viral gene pe38 is not only essential for the infectivity of CpGV but it is also the key factor in overcoming CpGV resistance in codling moth.The codling moth Cydia pomonella L. is a worldwide occurring insect pest that infests apples, pears, and walnuts. The larvae of codling moth bore into the fruit and cause severe economic damage if not controlled. A number of chemical and biological agents are available for the control of codling moth. One of the most efficient biological control agents (1–5) is the Cydia pomonella granulovirus (CpGV), which belongs to the dsDNA virus family Baculoviridae (genus Betabaculovirus). CpGV was first discovered in Mexico (Mexican isolate, CpGV-M) in 1963 (6); this isolate was later developed to commercial products now registered in 34 countries worldwide. The genome sequence of the in vivo cloned strain CpGV-M1 is ∼123 kbp and encodes for 143 ORFs (7). Based on SNPs in highly conserved genes, different geographic CpGV isolates were classified into four genome types, A–D (8). CpGV exhibits an extremely narrow host range that is restricted to C. pomonella and a very few closely related tortricids (Lepidoptera) (9).The development of resistance to baculovirus infection was thought to be unlikely before 2005 (10, 11), which is when the first cases of resistant codling moth populations with a 1,000- to 100,000-fold reduced susceptibility to commercial CpGV products containing the isolate CpGV-M were reported from organic apple plantations in Germany and France, where CpGV products had been intensively applied (12, 13). Since then, 38 apple plantations with CpGV resistance have been identified in Austria (2 orchards), Czech Republic (1), France (3), Germany (22), Italy (6), Switzerland (2), and the Netherlands (2) (14). For the codling moth strain CpR, which originated from a resistant field population in south Germany, as well as the genetically homogenous laboratory strain CpRR1, which derived from CpR, the mode of inheritance was revealed to be incompletely dominant, monogenic, and linked to the Z (sex) chromosome (15, 16). A similar mode of inheritance was also observed in resistant Czech and French codling moth populations arguing for a more or less universal mode of resistance in Europe (17, 18). These data led to the hypothesis that a genetic adaptation of codling moth to CpGV-M infection had occurred and was selected for by the intensive use of products containing CpGV-M (15).Insects manifest miscellaneous strategies to resist pathogens but lack an adaptive immune system. Insect defense to viral infections involves nonspecific factors such as physical barriers, enzymatic responses, and increasing ejection of infected midgut cells as larvae age, as well as specific factors involving cellular and humoral immunity (19). Physical barriers to infection include the perithropic membrane (PM), an ultrafilter for particles with size exclusion greater than 30 nm (20). Melanization of the cuticula mediated by phenoloxidase enzymes is involved in the encapsulation reaction of pathogens, such as bacteria, fungi, or virus-infected cells (21, 22). Developmental resistance, the decreasing susceptibility of larvae with increasing age, is rather common, and is mediated by infected midgut cell-sloughing (23–25). Behavior modifications also count among insect defense strategies (19). Feeding behaviors, e.g., can strongly affect the risk of insects for baculovirus infection as shown for Lymantria dispar L., which exhibit heritable cadaver-avoidance behaviors (26).An alternative way for insects to get rid of virus-infected cells is via programmed cell death—the apoptosis pathway (27). Baculoviruses, however, are able to block apoptosis with the help of different virus encoded antiapoptotic proteins. In Autographa californica multiple nucleopolyhedrovirus (AcMNPV), these proteins—namely P35 and P49—are inhibitors of the insect caspases. These inhibitors of apoptosis (IAPs), thereby, ensure a permissive virus infection (28).For codling moth, however, resistance based on the PM, the midgut, or the immune system has been excluded for CpGV (16, 29). Lack of CpGV DNA replication and a systemic resistance in all five instars indicated an early block to virus replication in resistant codling moth individuals (29, 30). At the same time, it was observed that certain CpGV isolates, such as I12 or NPP-R1/R4, were able to infect larvae from resistant codling moth strains (30, 31). Therefore, these naturally occurring CpGV isolates from different geographic origins were considered resistance-breaking isolates. Some of these isolates, meanwhile, replace CpGV-M in commercial biocontrol agents in Europe and demonstrate the importance of identifying resistance-breaking CpGV variants (30–32) for managing CpGV-resistant codling moth populations; however, their functional difference to CpGV-M allowing them to overcome CpGV resistance remained unknown.In this study we compared different naturally occurring geographic CpGV isolates representing all known CpGV genome types (8) in laboratory assays for their infectivity to susceptible (CpS) and resistant (CpRR1) codling moth strains. We found that all tested isolates—except CpGV-M—were able to overcome resistance in CpRR1 larvae. Whole-genome sequencing of these isolates revealed a single common difference in all resistance-breaking isolates, which was located in ORF24 (pe38). Therefore, occlusion bodies of bacmid-based recombinant CpGV (in the following termed pseudoviruses) knockout and recovery mutants of pe38 were established and tested for their activity in susceptible CpS and resistant CpRR1 larvae. The recovery of infectivity of a CpGV-M–based pseudovirus harboring the pe38 of the resistance-breaking CpGV-S in CpRR1 larvae demonstrated the key function of pe38 in overcoming baculovirus resistance in codling moth. In their entirety, these investigations demonstrate the significance of using multiple isolates of CpGV concurrently in field applications to avoid resistance in codling moth populations.     

A policy for leishmaniasis with respect to the prevention and control of drug resistance

At the moment no country has a policy designed to control or prevent drug resistance in leishmaniasis. The risk of resistance is high in areas of anthroponotic visceral leishmaniasis, for example North Bihar, India, where the rate in some areas is 60%. Post-epidemic Sudan is also at risk. Zoonotic areas in which HIV co-infection is common could also be at risk as sandflies can become infected from co-infected individuals. Many factors determine the choice of drug for the treatment of visceral leishmaniasis, and drug resistance may not be the over-riding priority. In anthroponotic areas reduction in transmission through public health measures will be important, but the use of two drugs in combination should be seriously considered. Pharmacokinetic and other features of the drugs available, relevant to their use in combination are discussed and tentative suggestions made concerning trials of possible combinations. These include miltefosine plus paromomycin and allopurinol plus an azole. Lessons may be learnt from the experiences of similar problems in malaria, leprosy and tuberculosis. Guidelines are offered for the introduction of policies to use drugs in combination, which differ between anthroponotic and zoonotic areas of transmission.     

Inhibited apoptosis and drug resistance in acute myeloid leukaemia

Despite extensive investigation into mechanisms of drug resistance in acute myeloid leukaemia (AML), the aetiology of therapeutic resistance is unclear. We found that five leukaemia cell lines (K562, HL-60, CEM, CEM induced to overexpress bcl-2, and REH) displayed parallel sensitivity to four antileukaemia drugs with different mechanisms of action, with K562 generally being the least sensitive and REH being the most sensitive. The amount of spontaneous apoptosis in the cell lines after serum-free culture paralleled their drug sensitivity: K562 cells displayed the least apoptosis at 24 h (2.50 ± 0.24%) and REH the most (24.47 ± 8.22%). The extent of spontaneous apoptosis of leukaemic blasts from 39 patients with newly diagnosed de novo AML also correlated with the success of the intensive, infusional cytarabine-based induction therapy. There was a median of 19.5% (range 3.6–64%) apoptotic AML cells after 24 h of serum-free culture in patients who entered a complete remission compared with 4.2% (1.8–7.0%) apoptotic AML cells in patients who did not achieve a complete remission ( P  = 0.0007). Thus, inhibited apoptosis was associated with both in vitro and in vivo pan-resistance to antileukaemic chemotherapy. The cause of inhibited apoptosis in AML is probably a function of interactions among multiple signals that influence apoptosis. Assessment of spontaneous apoptosis may serve as an important prognostic factor for AML.     

抗流感病毒药物和病毒耐药性

M2蛋白通道阻断剂金刚烷胺(amantadine)和金刚乙胺(rimantadine)在体内外均可迅速产生耐药和交叉耐药,机制是源于M2蛋白转膜区突变,涉及的5个主要氨基酸残基是26、27、30、31和34位.神经氨酸酶抑制剂磷酸奥司他韦(oseltamivir)、扎那米韦(zanamivir)和帕那米韦(parami...     

Increasing antimalarial drug resistance in Uganda and revision of the national drug policy

Chloroquine (CQ) resistance was first documented in Uganda in 1988. Subsequent surveillance of antimalarial drug resistance, conducted by the Ugandan Ministry of Health and several research organizations, suggests that resistance to CQ is now widespread, reaching critical levels in many areas of the country. In June 2000, the Ministry of Health held a National Consensus Meeting to evaluate the available drug efficacy data and review the national antimalarial drug policy. After extensive debate, the combination of CQ + sulfadoxine-pyrimethamine (SP) was chosen to replace CQ as the first-line treatment of uncomplicated malaria as an interim policy. This review evaluates the in vivo drug efficacy studies conducted in Uganda since 1988 and issues confronted in revision of the drug policy. The Ugandan experience illustrates the challenges faced by sub-Saharan African countries confronted with rising CQ resistance but limited data on potential alternative options. The choice of CQ + SP as a provisional policy in the absence of prerequisite efficacy, safety and cost-effectiveness data reflects the urgency of the malaria treatment problem, and growing pressure to adopt combination therapies. Surveillance of CQ + SP treatment efficacy, collection of additional data on alternative regimens and active consensus building among key partners in the malaria community will be necessary to develop a rational long-term antimalarial treatment policy in Uganda.     

HIV耐药评估和预防策略

HIV耐药会影响抗病毒治疗的长期疗效,使得艾滋病流行结束于2030年的目标难以实现。不同国家采用不同策略来预防和评估HIV耐药的产生和传播。目前,高收入国家对HIV感染者进行个体耐药检测。多数中低收入国家采用WHO提出的群体监测策略,监测耐药早期预警指标,评估治疗人群获得性耐药,监测治疗前耐药、传播性耐药及18月龄以下儿童耐药等措施。我国采用HIV耐药个体检测和群体监测相结合的防控策略。     

解脲脲原体生物群分布与耐药性差异研究

目的 了解本地区泌尿生殖道感染人群中解脲脲原体感染的生物群分布特点及其耐药情况。方法 采用液体选择培养基对泌尿道分泌物标本进行培养、鉴定及药敏;选取解脲脲原体阳性标本进行PCR分群和分型;分析各生物群型别及其耐药性差异。结果 985例检测标本中,单独解脲脲原体阳性395例,总阳性率为40.10%,其中女性368例,男性27例,其阳性率分别为42.45%、22.88%,女性阳性率高于男性,其差异有显著统计学意义(χ²=16.550,P<0.01)。395例单独解脲脲原体阳性标本作PCR分群分型显示,单独生物一群阳性278例,阳性率为70.89%;单独生物二群阳性80例,阳性率为20.25%。单独生物一群中以单一血清6型(121例,43.21%)、3/14型(96例,35.53%)感染为主。单独生物二群均为混合血清型感染,基因亚型1(血清型2、5、8、9)和亚型3(血清型7、11)混合感染(49例,61.25%);基因亚型2(血清型4、10、12、13)和亚型3(血清型7、11)混合感染共检出(31例,38.75%)。结合药敏结果发现,解脲脲原体对喹诺酮类药物耐药率较高,对四环素类药物较敏感,且多重耐药现象严重。生物一群对司帕沙星的耐药性高于生物二群,二者差异有统计学意义(χ²=5.701,P=0.017<0.05),且生物一群对米诺环素、多西环素、阿奇霉素的耐药性远高于生物二群,其差异有显著统计学意义(χ²=10.811,χ²=19.103,χ²=7.541,P均<0.01)。进一步结合分型结果发现,生物一群中单一血清型S6耐药率最高的抗菌药为氧氟沙星(76.86%),S1型耐药率最高的抗菌药为罗红霉素(95.74%),S3/14型耐药率最高的抗菌药为罗红霉素和阿奇霉素(均为86.46%)。结论 泌尿生殖道感染人群中解脲脲原体阳性率较高,女性高于男性,以生物一群单一血清型感染为主;本地区耐药形势严峻,生物一群耐药性高于生物二群;生物一群中各血清型的耐药性存在不同程度的差异。结合解脲脲原体生物分群分型结果可辅助合理选用抗菌药物,以避免滥用致耐药株产生。     

慢性阻塞性肺部疾病急性加重期病原菌分布及耐药性研究

目的探讨慢性阻塞性肺疾病急性加重期(AECOPD)的病原菌分布和耐药性。方法收集柳州多家医院2012年2月至2014年6月期间呼吸内科的AECOPD患者805例,详细记录入组患者的临床资料,分析AECOPD患者的病原菌分布及耐药特点。结果 805例AECOPD患者中分离出病原菌482例,共分离病原菌602株,其中革兰氏阴性菌430株,占71.5%,革兰氏阳性菌97株,占16.1%,真菌75株,占12.5%;铜绿假单胞菌、肺炎克雷伯菌、大肠埃希菌居前3位,分别占20.3%、18.3%和13.8%;肺炎克雷伯菌和大肠埃希菌对哌拉西林耐药率均80.0%,大肠埃希菌对头孢哌酮、头孢曲松、环丙沙星、左氧氟沙星耐药率较高,均50.0%。结论 AECOPD病原菌以革兰阴性杆菌为主,治疗应根据病原菌及药敏试验结果指导,临床合理选用抗菌药物。     

猪链球菌9型分离株的耐药性及耐药基因分析

目的 了解猪链球菌9型(Streptococcus suis serotype 9,SS9)的耐药性和耐药基因情况。方法 采用药敏纸片扩散法检测15株SS9菌株对9类21种不同抗生素的耐药性,并通过PCR方法检测其大环内酯类及四环素类耐药基因的携带情况。结果 被检菌株对大环内酯类及林可胺类耐药率都为100%,对四环素及链霉素的耐药率都为93.3%,对头孢类、氯霉素类、阿莫西林、糖肽类以及庆大霉素较为敏感。所有菌株均耐7种及以上抗菌药物,最高可耐16种,其中以8重耐药的菌株数量最多。不同地区SS9分离株对抗生素耐药情况不同,健康猪分离株耐药情况较病猪分离株更为严重。100%的菌株具有ermB基因,93.33%的菌株具有tetO基因,表明ermB和tetO分别可能是介导SS9对大环内酯类及四环素产生耐药性的主要原因之一。结论 本研究为猪链球菌9型的预防和临床治疗、耐药性及耐药机制的研究提供了参考依据。