Sterile Blood Research Articles

In Vitro and in Vivo Evaluation of Antibiotic Combination against Multidrug Resistant Enterobacter Species Isolated From Patients of a Tertiary Care Hospital, Bangladesh

The emergence of multidrug-resistant (MDR) Enterobacter as a worrying resistant pathogen is an important health concern, especially when there is scarcity of new antibiotics active against Gram-negative bacteria. However, currently no defined therapies available for MDR Enterobacter infections. In this study, in vitro and in vivo efficacy of different antimicrobial combinations were assessed. This cross-sectional study was carried out in the department of Microbiology of Dhaka medical college hospital, Bangladesh from July, 2018 to June, 2019. Multidrug resistance among isolated Enterobacter species were detected phenotypically by disk diffusion method. PCR and sequencing of fosfomycin resistance genes were done. In vitro activity of fosfomycin, amikacin, imipenem, piperacillin-tazobactam and their combinations were evaluated using agar dilution method and synergy was assessed by Fractional inhibitory concentration index. Mice models were made by using the MDR Enterobacter strain. We evaluate the efficacy of fosfomycin, amikacin, imipenem and their combination against multi-drug resistant Enterobacter infection in experimental mice models. Among 28 isolated Enterobacter spp. 53.33% were multidrug-resistant. Among the fosfomycin resistant Enterobacter spp. 70% , 50%, 40% were positive for fosA. foA5 and fosB2 respectively. The fractional inhibitory concentration index indicated that combining antibiotics resulted 2 to 8 fold reduction of MIC compared to single therapy. The ratio of synergy observed in imipenem-amikacin, fosfomycin-amikacin, fosfomycin-imipenem 16.67%, 87.33%, 50.0% respectively in vitro. No synergy observed in imipenem-piperacillin tazobactam combination. In mice model, compared to single antibiotic therapy, fosfomycin-amikacin, imipenem-amikacin, fosfomycin-imipenem showed increased sterile blood culture (100%, 60%, 80%). Fosfomycin plus amikacin or fosfomycin plus imipenem may be alternative treatment option against multidrug-resistant Enterobacter infection.

A simple, efficient and cost-effective method for medium- to longterm maintenance and storage of Mycobacterium abscessus complex organisms

Traditional culture of non-tuberculous mycobacteria (NTMs) has involved egg-based formulations (Lowenstein-Jensen medium, Ogawa Egg medium) or defined media (Middlebrook formulations), which have disadvatages of composition complexity, availability and cost. Previously, the commercial agar formulation, Standard Plate Count (SPC) agar [Yeast extract 2.5 g/L, pancreatic digest of casein 5.0 g/L, glucose 1.0 g/L, agar 15.0 g/L, pH 7.0 ± 0.2 at 25 °C] has been shown to be an effective solid medium for the enumeration and laboratory manipulation of Mycobacterium abscessus complex organisms. Given its relative simplicity, commercial availability and inexpensive cost, we wished to evaluate its utility for the medium- to longterm maintenance/storage of these organisms. M. abscessus complex organisms (n = 33), were inoculated onto SPCA slopes and stored undistubed in the dark at ambient temperature for six months. Following this, slopes were broken out and culture of the NTM attempted. All slopes maintained NTM culture viability and were able to initiate growth six months later. We therefore advocate the cost-effective employment of SPCA slopes for the medium- to longterm maintenance of M. abscessus organisms, without the need for complex media, availability of sterile blood and requirements for continuous -80 °C freezing.

How to Handle CT-Guided Abscess Drainages in Microbiological Analyses? Sterile Vials vs. Blood Culture Bottles for Transport and Processing.

The aim of this investigation was to compare microbiological analyses of 100 computed tomography-guided drainages from infectious foci (thoracic, abdominal, musculoskeletal), transported and analyzed by two widely established techniques, that are (i) sterile vials or (ii) inoculated blood culture bottles. The mean number of detected microorganisms from blood culture (aerobic/anaerobic) or conventional method (sterile vial, solid and broth media) per specimen were comparable with 1.29 and 1.41, respectively (p = 1.0). The conventional method showed a trend towards shorter time-to-result (median 28.62 h) in comparison to blood culture incubation (median 43.55 h) (p = 0.0722). Of note, detection of anaerobes (13% vs. 36%) and the number of detected microorganisms in polymicrobial infections (2.76 vs. 3.26) differed significantly with an advantage towards conventional techniques (p = 0.0015; p = 0.035), especially in abdominal aspirations. Despite substantially overlapping results from both techniques, the conventional approach includes some benefits which justify its role as standard approach.

Incidence and Effects of Acquisition of the Phage-Encoded ssa Superantigen Gene in Invasive Group A Streptococcus.

The acquisition of the phage-encoded superantigen ssa by scarlet fever-associated group A Streptococcus (Streptococcus pyogenes, GAS) is found in North Asia. Nonetheless, the impact of acquiring ssa by GAS in invasive infections is unclear. This study initially analyzed the prevalence of ssa+ GAS among isolates from sterile tissues and blood. Among 220 isolates in northern Taiwan, the prevalence of ssa+ isolates increased from 1.5% in 2008–2010 to 40% in 2017–2019. Spontaneous mutations in covR/covS, which result in the functional loss of capacity to phosphorylate CovR, are frequently recovered from GAS invasive infection cases. Consistent with this, Phostag western blot results indicated that among the invasive infection isolates studied, 10% of the ssa+ isolates lacked detectable phosphorylated CovR. Transcription of ssa is upregulated in the covS mutant. Furthermore, in emm1 and emm12 covS mutants, ssa deletion significantly reduced their capacity to grow in human whole blood. Finally, this study showed that the ssa gene could be transferred from emm12-type isolates to the emm1-type wild-type strain and covS mutants through phage infection and lysogenic conversion. As the prevalence of ssa+ isolates increased significantly, the role of streptococcal superantigen in GAS pathogenesis, particularly in invasive covR/covS mutants, should be further analyzed.

Improved diagnostic prediction of the pathogenicity of bloodstream isolates of Staphylococcus epidermidis.

With an estimated 440,000 active cases occurring each year, medical device associated infections pose a significant burden on the US healthcare system, costing about $9.8 billion in 2013. Staphylococcus epidermidis is the most common cause of these device-associated infections, which typically involve isolates that are multi-drug resistant and possess multiple virulence factors. S. epidermidis is also frequently a benign contaminant of otherwise sterile blood cultures. Therefore, tests that distinguish pathogenic from non-pathogenic isolates would improve the accuracy of diagnosis and prevent overuse/misuse of antibiotics. Attempts to use multi-locus sequence typing (MLST) with machine learning for this purpose had poor accuracy (~73%). In this study we sought to improve the diagnostic accuracy of predicting pathogenicity by focusing on phenotypic markers (i.e., antibiotic resistance, growth fitness in human plasma, and biofilm forming capacity) and the presence of specific virulence genes (i.e., mecA, ses1, and sdrF). Commensal isolates from healthy individuals (n = 23), blood culture contaminants (n = 21), and pathogenic isolates considered true bacteremia (n = 54) were used. Multiple machine learning approaches were applied to characterize strains as pathogenic vs non-pathogenic. The combination of phenotypic markers and virulence genes improved the diagnostic accuracy to 82.4% (sensitivity: 84.9% and specificity: 80.9%). Oxacillin resistance was the most important variable followed by growth rate in plasma. This work shows promise for the addition of phenotypic testing in clinical diagnostic applications.

Qualitative Evaluation of Platelet Concentrates Prepared with Different Methods

Introduction: One of the most important responsibility of Blood Bank is to ensure uninterrupted supply of quality blood products, namely Platelet Concentrate (PC), which are in high demand particularly during pandemic. In India, PCs are of three types prepared by different methods like Platelet Rich Plasma-Platelet Concentrate (PRP-PC), Buffy Coat Platelet Concentrates (BC- PC), and Single Donor Platelets (SDP) units. Hence, establishing optimum quality standards across all the three types of PC remains a challenge in order to maximum benefit to recipients. Aim: To assess and compare the quality parameters of platelet concentrate prepared by PRP-PC, BC-PC and apheresis methods (SDP). Materials and Methods: This laboratory based observational study was conducted at rural, tertiary care, postgraduate, teaching hospital located at South India from December 2018 to November 2019. A total of 156 PCs were studied with 52 units belonging to PRP method, 52 units BC-PC method and the remaining were 52 units of SDP prepared by apheresis method. Quality parameters include volume, pH, swirling, White Blood Cell (WBC) count/bag, platelet count/bag and platelet indices. Platelet indices include Mean Platelet Volume (MPV), Platelet Distribution Width (PDW), and Platelet Large Cell Ratio (PLCR). Arterial Blood Gas (ABG) analysis was also done to assess (pH, partial pressure of Carbon Dioxide (pCO2), plasma bicarbonate (pHCO3-)). The p-value less than 0.05 were considered statistically significant. Statistical Package for the Social Sciences (SPSS) software version 23.0 used for statistical analysis. Results: All the 156 PC units were analysed as per the Director General of Health Services (DGHS) India, criteria. Among the 156 platelet concentrates scored as per DGHS criteria, a score of 5 was obtained for 34.61% [18/52], 30.76 [16/52] and 11.5% [6/52] SDP, BC-PC and PRP-PC units, respectively. All the PC units were sterile and Red Blood Cell (RBC) contamination was within acceptable limits. Conclusion: In this study, quality parameters of SDP was better than BC-PC and PRP-PC but with improvement and more standardisation of procedures and storage, BC-PC units can give a yield similar to that of SDP units.

EVALUATION OF EFFECT OF THE STORAGE TIME ON PLATELET CONCENTRATION IN PLATELET RICH FIBRIN- BIOCHEMICAL AND MICROSCOPIC ANALYSIS

Aim of the present study was to evaluate the effect of storage time on platelet-rich fibrin (PRF) and the stability of the fibrin clot over a period of time. Three blood samples were drawn from participants in sterile blood sampling tube. Two blood samples drawn in sterile glass tubes were centrifuged to form PRF. Third non-centrifuged sample drawn in EDTA-containing tube was used to measure baseline blood platelet concentration. After PRF had formed, it was removed from respective test tubes at different time intervals i.e. immediately after centrifugation- PRF A (Group A) and after 60 min of storage time in the blood collecting tube- PRF B (Group B). Residual blood from each group was tested for platelet concentration and compared with baseline reading. Similarly, PRF membranes were studied microscopically immediately after centrifugation and other after 60 minutes. Platelet concentration of PRF in blood and PRF membrane for both groups was calculated using the difference between baseline and residual platelet concentration- biochemically and microscopically. A paired t-test showed a statistically significant difference between the two groups (p<0.005).

Role of Hand Hygiene and Sterile Blood Sampling in Reduction of False Positive Blood Cultures from a Neonatal Intensive Care Unit of a Rural Teaching Hospital: A Quasi-experimental Study

Introduction: Blood culture remains the gold standard for diagnosis of newborn infections. Its contamination not only results in inadvertent use of antimicrobials against the false positives but also causes substantial financial burden and patient safety concerns. There are various ways to reduce blood contamination, of which hand hygiene and sterile blood sampling are most crucial. Aim: To evaluate and reduce the blood contamination rate and false positive blood cultures from the Neonatal Intensive Care Unit (NICU). Materials and Methods: The prevalent bacteriology and Blood Culture Contamination Rate (BCCR) at the NICU was assessed by a retrospective study on all the neonates with suspected sepsis who had blood culture at admission. Healthcare workers at NICU were observed for their hand wash and sterile blood sampling technique by dedicated team members. Identified deficiencies among them were intervened by an educational model with regular training and reinforcement on hygienic practices for three months. Postintervention prospective study was done on all the neonates with suspected sepsis who had blood culture at admission. During this period the same educational model continued, to know its efficacy in reduction of blood contamination and change of bacteriology pattern. Data collected was entered into Microsoft excel sheet 2010 version and analysed using software Stata 14.0 version. Results: Preintervention blood culture reports of 205 neonates screened for sepsis showed 58 (28.3%) samples with positive growth. Of this, 18 were contaminants, 31 were Coagulase Negative Staphylococci (CoNS), three gram negative and six gram positive organisms with blood contamination rate of 8.7%. Blood culture reports of postintervention study on 200 neonates screened for sepsis showed 41 (20.5%) samples positive for growth. Bacteriology showed 12 contaminants, 13 CoNS, eight gram positives, seven gram negatives and one fungal growth with BCCR of 6%. After intervention growth of CoNS was reduced which was statistically significant (p<0.05). Conclusion: Simple inexpensive education programme helps the healthcare workers to follow the standard technique of hand wash and sterile blood sampling which goes a long way in reducing blood culture contamination.

Corynebacterium striatum Cardiac Device-Related Infective Endocarditis: The First Case Report in a Patient With a Cardiac Resynchronization Therapy Defibrillator Device and Review of the Literature

Corynebacterium striatum (C. striatum) is a skin commensal agent, rarely described as a cause of infective endocarditis. We describe a case of a 48-year-old man, with multiple comorbidities with cardiac resynchronization therapy defibrillator (CRT-D) device implanted 1 year before. A cardiac device-related infective endocarditis (CDRIE) due to C. striatum, with vegetations in the tricuspid valve adjacent to the electrode lead and concomitant lumbar spondylodiscitis were diagnosed. The patient was treated initially with a 6-week course of vancomycin with sterile blood cultures and reduction of inflammatory parameters. Surgery was refused at this stage. Six weeks later, he was readmitted due to C. striatum bacteriemia recurrence, with vegetations adhering to the electrode wire, being treated with daptomycin 10mg/kg body weight, after presenting renal toxicity to vancomycin. CRT-D device was removed with implantation of epicardial cardiac resynchronization therapy pacemaker (CRT-P). To our knowledge, this might be the first description of C. striatum CDRIE in a patient with a CRT-D. In the five cases described in the literature of CDRIE by this agent, early removal of the pacemaker was performed with good results. In this case, the device was removed only after failure of medical treatment alone.

Blood culture contamination rates at different level healthcare institutions in the Western Cape, South Africa.

Sterile blood culture (BC) collection procedures are important to prevent the consequences of contamination, namely, prolonged patient hospitalisation, unnecessary antimicrobial exposure and an increase in hospital costs. Blood culture contamination rates were determined at different hospitals in the Cape Metropole over a 3-year period. Study findings showed that contaminated BCs have a financial impact on the healthcare system and contamination rates remain above accepted international standards, except in the presence of a phlebotomist team. High BC contamination rates might be reduced by the implementation of cost-effective educational intervention programmes, which reminds healthcare workers to collect BC samples aseptically.

Glossary

Glossary

Proof of Concept of Culturomics Use of Time of Care.

Culturomics, a high throughput culture method with rapid identification of the colonies by Matrix Assisted Laser Desorption Ionization/Time Of Flight Mass Spectrometry (MALDI-TOF MS), has demonstrated its contribution to the exploration of the gut microbiota over the past 10 years. However, the cost, work time and workload, considerably limit its use on a large scale or emergency context. Here, by testing two different stool samples, including a stool sample from a patient requiring rapid immunotherapy treatment, we tested a new fast culturomic protocol using two pre-incubation media, blood culture bottle and YCFA modified medium. Both media were supplemented with 2 ml of rumen fluid filtered at 0.2 μm and 2 ml of defibrinated and sterile sheep blood. Unlike the standard culturomics, subculturing of blood culture bottle were performed at reduced incubation time (3 h, 6 h, 9 h, 24 h) and at a longer incubation time (3 days, 7 days, and 10 days) at 37°C. By testing 5,200 colonies per MALDI-TOF MS and obtaining a comparable number of cultured bacterial species (131 to 143) in a stool sample, this new protocol reduced the number of colonies tested by 57%, working time by 78.6% and cost by 72.2%. In addition, we highlighted that the proportion of strict anaerobic species has increased by 24%, known to be the preferential targets for biotherapy, including Faecalibacterium prausnitzii, Akkermansia muciniphila, Christensenella minuta, and Phascolarctobacterium faecium. Finally, this work showed that some bacterial species grew earlier but disappeared with prolonged incubation times.

Adherence of Streptococcus mutans to Nanoceramic and Nanohybrid Resin Composites: An In vitro Study

Streptococcus mutans (S. mutans) is one of the cariogenic microorganisms. The restorative materials which harbor a biofilm with high number of S. mutans can accelerate the occurrence of dental caries. The purpose of this study was to evaluate the adherence of S. mutans to nanoceramic and nanohybrid resin composites. Fifteen discs of each material (Nanohybrid resin composite, Nanoceramic resin composite) were prepared, polished, and sterilized in a gamma radiation chamber. Specimens were exposed to the S. mutans bacterial suspension (0.5 McFarland) and were incubated for 4 hours. Specimens were rinsed and sonicated in normal saline, 10 μl of the obtained suspension was cultured in a sterile blood agar medium. After 24 hours, the number of colony forming units of S. mutans was counted. A sterility test control was considered for each group of materials. The data was analyzed by Independent t test. The means and standard deviations of the logarithmic counts of the colonies on the surfaces of nanohybrid resin composites and nanoceramic resin composite were equal to 3.2±0.87 and 2.8±0.324 respectively. Ceram X Universal did not show any significant difference in the bacterial adhesion compared to Filtek Z350XT. Both composites showed similar behaviour in terms of S. mutans colonization in a simple biofilm formation model.

Trimethoprim-sulfamethoxazole Versus Azithromycin for the Treatment of Undifferentiated Febrile Illness in Nepal: A Double-blind, Randomized, Placebo-controlled Trial.

BackgroundAzithromycin and trimethoprim-sulfamethoxazole (SXT) are widely used to treat undifferentiated febrile illness (UFI). We hypothesized that azithromycin is superior to SXT for UFI treatment, but the drugs are noninferior to each other for culture-confirmed enteric fever treatment.MethodsWe conducted a double-blind, randomized, placebo-controlled trial of azithromycin (20 mg/kg/day) or SXT (trimethoprim 10 mg/kg/day plus sulfamethoxazole 50 mg/kg/day) orally for 7 days for UFI treatment in Nepal. We enrolled patients >2 years and <65 years of age presenting to 2 Kathmandu hospitals with temperature ≥38.0°C for ≥4 days without localizing signs. The primary endpoint was fever clearance time (FCT); secondary endpoints were treatment failure and adverse events. ResultsFrom June 2016 to May 2019, we randomized 326 participants (163 in each arm); 87 (26.7%) had blood culture–confirmed enteric fever. In all participants, the median FCT was 2.7 days (95% confidence interval [CI], 2.6–3.3 days) in the SXT arm and 2.1 days (95% CI, 1.6–3.2 days) in the azithromycin arm (hazard ratio [HR], 1.25 [95% CI, .99–1.58]; P = .059). The HR of treatment failures by 28 days between azithromycin and SXT was 0.62 (95% CI, .37–1.05; P = .073). Planned subgroup analysis showed that azithromycin resulted in faster FCT in those with sterile blood cultures and fewer relapses in culture-confirmed enteric fever. Nausea, vomiting, constipation, and headache were more common in the SXT arm.ConclusionsDespite similar FCT and treatment failure in the 2 arms, significantly fewer complications and relapses make azithromycin a better choice for empirical treatment of UFI in Nepal.Clinical Trials RegistrationNCT02773407.

Transcriptomic profiling of the digestive tract of the rat flea, Xenopsylla cheopis, following blood feeding and infection with Yersinia pestis.

Yersinia pestis, the causative agent of plague, is a highly lethal pathogen transmitted by the bite of infected fleas. Once ingested by a flea, Y. pestis establish a replicative niche in the gut and produce a biofilm that promotes foregut colonization and transmission. The rat flea Xenopsylla cheopis is an important vector to several zoonotic bacterial pathogens including Y. pestis. Some fleas naturally clear themselves of infection; however, the physiological and immunological mechanisms by which this occurs are largely uncharacterized. To address this, RNA was extracted, sequenced, and distinct transcript profiles were assembled de novo from X. cheopis digestive tracts isolated from fleas that were either: 1) not fed for 5 days; 2) fed sterile blood; or 3) fed blood containing ~5x108 CFU/ml Y. pestis KIM6+. Analysis and comparison of the transcript profiles resulted in identification of 23 annotated (and 11 unknown or uncharacterized) digestive tract transcripts that comprise the early transcriptional response of the rat flea gut to infection with Y. pestis. The data indicate that production of antimicrobial peptides regulated by the immune-deficiency pathway (IMD) is the primary flea immune response to infection with Y. pestis. The remaining infection-responsive transcripts, not obviously associated with the immune response, were involved in at least one of 3 physiological themes: 1) alterations to chemosensation and gut peristalsis; 2) modification of digestion and metabolism; and 3) production of chitin-binding proteins (peritrophins). Despite producing several peritrophin transcripts shortly after feeding, including a subset that were infection-responsive, no thick peritrophic membrane was detectable by histochemistry or electron microscopy of rat flea guts for the first 24 hours following blood-feeding. Here we discuss the physiological implications of rat flea infection-responsive transcripts, the function of X. cheopis peritrophins, and the mechanisms by which Y. pestis may be cleared from the flea gut.

Evaluation of Two Methods for the Detection of Third Generation Cephalosporins Resistant Enterobacterales Directly From Positive Blood Cultures.

Due to the importance of a rapid determination of patients infected by multidrug resistant bacteria, we evaluated two rapid diagnostic tests for the detection of third-generation cephalosporins (3GC)-resistant Enterobacterales directly from positive blood cultures within 1 h: BL-REDTM (electrochemical method) and β-LACTATM test (chromogenic method). A panel of 150 clinical strains characterized for their resistance profiles (e.g., penicillinases, extended-spectrum beta-lactamases (ESBLs), overproduction of cephalosporinase, carbapenemases, impermeability) was tested. Approximately 100 CFU of each isolate was spiked into sterile blood culture bottles and incubated in a BD BACTECTM FX automated system (Becton Dickinson, USA). Positive blood cultures were examined to parallel testing using the BL-REDTM and β-LACTATM tests and conventional susceptibility method (disc diffusion following EUCAST recommendations). For all phenotypes combined, the sensitivity, specificity, positive predictive value, and negative predictive value in the detection of 3GC resistance were, respectively (i) with BL-REDTM: 45.7, 100, 100, and 54.2% and (ii) with β-LACTATM test: 52.2, 100, 100, and 56.9%. The positivity of tests allows to adapt antibiotic treatment whereas the negative result requires other tests. Moreover, these tests detect most Ambler class A-producing Enterobacterales (KPC, ESBL, extended-spectrum OXY) with sensitivities and specificities of 87.5 and 99% for BL-REDTM, respectively and both 100% for β-LACTATM test (47/47 isolates). These two rapid tests failed to detect AmpC overexpressed (sensitivities of 2.7% for BL-REDTM and 0% for β-LACTATM test) and Ambler class B-producing Enterobacterales (sensitivities of 40% for both tests) notably strains without ESBLs associated (sensitivities of 0% for both tests). BL-REDTM and β-LACTATM tests are easy-to-use and mainly attractive when a positive result is obtained notably to detect most of the Ambler class A-producing Enterobacterales in <1 h after the positivity of the blood culture, allowing a rapid adaptation of the antibiotic therapy in patients.

Interleukin-6 versus C-reactive protein as markers for early detection of bacteremia in febrile neutropenia in pediatric population

Abstract Introduction: Systemic infection leading to multiorgan failure during neutropenia is one of the leading causes of treatment-related mortality among children receiving chemotherapy. Reliable markers are needed to diagnose or rule out infection, to reduce the empirical use of broad-spectrum antibiotic therapy. Aims and Objectives: The aim is to compare the role of interleukin-6 versus C-reactive protein (CRP) as markers of sepsis in febrile neutropenia in pediatric patients while on chemotherapy for malignancy. Materials and Methods: This was a prospective observational study carried out in the Department of Paediatrics of a tertiary care Hospital in South India. All children with malignancy in the age group from 1 month to 18 years diagnosed to have febrile neutropenia during any phase of chemotherapy were included in the study. Multiple episodes of febrile neutropenia in the same child were analyzed as separate episodes. Results: Thirty-two episodes of febrile neutropenia were analyzed. There were 7 microbiologically documented infections (MDI), 19 clinically documented infections and 6 episodes of fever of unknown origin. Out of the 7 MDI, 5 were Gram-negative sepsis and 2 were Gram-positive sepsis. Gram-negative sepsis had a much higher median IL-6 value (169) than Gram-positive sepsis (17.5) and sterile blood cultures (52). However, median value of CRP was only slightly higher in Gram-positive sepsis (85.5) than in Gram-negative sepsis (60.7) and sterile blood cultures (44.2). Conclusion: In this study, higher IL-6 values predicted Gram-negative sepsis better than CRP.

Acute kidney injury with dark urine: the case of paroxysmal nocturnal hemoglobinuria

In this case report; we describe a patient with severe attack of paroxysmal nocturnal hemoglobinuria (PNH) following Ciprofloxacin therapy. He presented with recurrent abdominal pain, repeated vomiting and dark urine. Physical examination revealed severe pallor and jaundice. Laboratory investigations showed severe intravascular hemolysis with negative Coomb’s test, progressive acute kidney injury (AKI), sterile blood cultures and negative serology for autoimmune diseases. Subsequently, he developed pancytopenia. Ham test was positive and flow cytometry, later on, confirmed PNH. He was supported with multiple transfusions of packed blood cells and hemodialysis. Ciprofloxacin was discontinued and his PNH was treated with Solumedrol 1 g daily for 3 days followed by Prednisone 60 mg/day for 2 months. Two weeks later; his hemolysis abated and his AKI improved. Up to 5 years later, he still has minor PNH clone yet without disease activity. In conclusion; our patient had acute drug-induced hemolytic crisis associated with minor PNH clone. With drug-vigilance; no further relapses were reported and his PNH clone remained stable for 5 years. The case expands the spectrum of PNH phenotypes and its triggering factors.&#x0D; Keywords: AKI, anemia, autoimmune, flow cytometry, hemolysis, PNH.

Estimation of life expectancy and measurement of immature stages ofLucilia sericatafed on three kinds of diets

Background and Objectives: Finding the best diet is very important to rear Lucilia sericata larvae for therapeutic purposes and prepare standard curves in forensic entomology. The aim of this study was to find the best diet for larvae in maggot therapy. Furthermore, this study was conducted to obtain a vertical life table and measure the length and width of immature stages of L. sericata for forensic entomology. Materials and Methods: Larvae of L. sericata (Karaj strain) were used to evaluate diets. The tests were carried out in three replicates of 100 eggs for each diet including chicken liver, blood agar, and fish food at the same time. Independent t-test, ANOVA, and Tukey’s post-hoc tests were used to compare the mean length and width of larvae between different groups. A P-value of less than 0.05 was considered significance level. Results: In contrast to the first and the second larval instars, there was a significant difference in the mean length and width of the third instar larvae (P&lt;0.001). At the third instar larval stage, those larvae that had been fed on chicken liver were significantly larger than the others. There were also no significant differences between life expectancy, overall survivorship rate, and force of mortality of larval groups fed on three different diets. Conclusions: Chicken liver is an advisable diet for nurturing larvae and plotting standard curves in forensic entomology. For rearing the first and the second larval instars in sterile conditions, blood agar and fish food can serve as suitable diets.

In vitro and in vivo Evaluation of Antibiotic Combination against Imipenem Resistant Acinetobacter baumannii Strains Isolated from Bangladeshi Patients

Acinetobacter baumannii is becoming a common etiological agent of nosocomial infections resulting in septicemia, meningitis, endocarditis, pneumonia, wound, and urinary tract infections within the healthcare setting. There are currently no defined optimal therapies available for multidrug-resistant (MDR) Acinetobacter baumannii infections. We evaluated the efficacy of imipenem, ceftazidime, amikacin and their combinations against imipenem resistant A. baumannii in experimental rat models. We also detected MBL encoding genes such as blaNDM-1, blaVIM and blaIMP and ESBL encoding genes such as blaCTX-M-15 and blaOXA-1 genes by Polymerase chain reaction (PCR). MBL encoding genes such as blaNDM-1 (83.33%), blaVIM (66.67%) and blaIMP (41.67%) and ESBL encoding genes such as blaCTX-M-15 (16.67%) and blaOXA-1 (12.50%) were detected among imipenem resistant Acinetobacter baumannii by PCR. In vitro activities of imipenem, ceftazidime, amikacin, ciprofloxacin, tigecycline and their combinations were tested using agar dilution method. The proportions of synergy observed in imipenem-ceftazidime, imipenem-amikacin, imipenem-ciprofloxacin and amikacin-tigecycline combinations were 25%, 54.17%, 12.50% and 41.67% respectively in vitro. Rat septicaemic models were evaluated using the imipenem resistant A. baumannii strain. Rats were treated with three antimicrobials and their combinations. For the rat model, the efficacies of imipenem, amikacin and imipenem plus ceftazidime and imipenem plus amikacin were assayed. In the septicaemic rat model, compared to the control group, (i) imipenem alone, (ii) ceftazidime alone (iii) imipenem plus ceftazidime (iv) imipenem plus amikacin showed increased sterile blood culture (50%, 16.67%, 66.67% and 83.33%). Imipenem plus amikacin or amikacin plus tigecycline may be appropriate for the treatment against imipenem resistant Acinetobacter baumannii infections.