Performance of an in-house real-time PCR assay for detecting Cytomegalovirus infection among transplant patients from a tertiary care centre.

Background: Quantitative Cytomegalovirus (CMV) polymerase chain reactions are increasingly being used for monitoring CMV DNAemia in haematopoietic stem cell transplants and solid organ transplants. Objective: In this study, a commercial CMV viral load assay was compared with an in-house viral load assay. Materials and Methods: A total of 176 whole-blood samples were tested for CMV DNAemia using both assays. Results: Our evaluation showed a difference of 1 log10copies/ml between the two assay systems in determining CMV viral loads in the clinical samples. Conclusion: The in-house viral load assay had a better correlation with clinical findings compared to the commercial assay. Quality assessment of these assays was done by the United Kingdom National External Quality Assessment Scheme (UKNEQAS), an external proficiency testing programme, and by the National Institute for Biological Standard and Control (NIBSC) standard. For UKNEQAS and NIBSC standards, the bias between the assays was 0.73 log10and 0.85 log10, respectively. This difference is well within the acceptable range already reported in the literature.

Quantification of live Lactobacillus acidophilus in mixed populations of live and killed by application of attenuated reflection Fourier transform infrared spectroscopy combined with chemometrics.

Since culture-based methods are costly and time consuming, alternative methods are investigated for the quantification of probiotics in commercial products. In this work ATR- FTIR vibration spectroscopy was applied for the differentiation and quantification of live Lactobacillus (La 5) in mixed populations of live and killed La 5, in the absence and in the presence of enteric polymer Eudragit® L 100-55. Suspensions of live (La 5_L) and killed in acidic environment bacillus (La 5_K) were prepared and binary mixtures of different percentages were used to grow cell cultures for colony counting and spectral analysis. The increase in the number of colonies with added%La 5_L to the mixture was log-linear (r2 = 0.926). Differentiation of La 5_L from La 5_K was possible directly from the peak area at 1635 cm-1 (amides of proteins and peptides) and a linear relationship between%La 5_L and peak area in the range 0-95% was obtained. Application of partial least squares regression (PLSR) gave reasonable prediction of%La 5_L (RMSEp = 6.48) in binary mixtures of live and killed La 5 but poor prediction (RMSEp = 11.75) when polymer was added to the La 5 mixture. Application of artificial neural networks (ANNs) improved greatly the predictive ability for%La 5_L both in the absence and in the presence of polymer (RMSEp = 8.11 × 10-8 for La 5 only mixtures and RMSEp = 8.77 × 10-8 with added polymer) due to their ability to express in the calibration models more hidden spectral information than PLSR.

Mycobacterium chelonae causing chronic wound infection and abdominal incisional hernia.

Mycobacterium chelonae is a rapidly growing mycobacterium that is found all over the environment, including sewage and tap water. They are important species associated with chronic non-healing wounds. We report a case in a 41 year old female patient who underwent multiple surgeries for an ovarian cyst, tubo-ovarian abscesses with peritonitis and a repair of an abdominal incisional hernia.

First Report of OXA-4, an ESBL Isolated from Pseudomonas aeruginosa a South Indian Strain.

The OXA-type ß-lactamases are so named because of their oxacillin-hydrolyzing abilities. In this study we characterize an extended spectrum ß-lactamase, designated OXA-4, produced by a clinical isolate of Pseudomonas aeruginosa. ESBL production was detected by double disk synergy test. The P. aeruginosa isolate was obtained from endotracheal suction tip of 84 years old male patient diagnosed with CVA and hypertension. ESBL producing OXA ß-lactamases was detected by PCR with primers specific to the conserved regions of the coding genes. Iso electric focusing was done to confirm the significance, sequencing the amplified product was also done. In the phenotypic identification, the strain was highly resistant to third generation cephalosporins and also to imipenem. The PCR amplified product for OXA ß-lactamase was viewed at 919 bp. The pI point for the same was identified at 7.2. With the help of sequencing the amplified OXA ß-lactamase was identified as OXA-4 gene. Here we report P. aeruginosa producing OXA-4 ESBL for the first time in the Indian subcontinent.

International evidence-based recommendations on ultrasound-guided vascular access.

PURPOSE: To provide clinicians with an evidence-based overview of all topics related to ultrasound vascular access. METHODS: An international evidence-based consensus provided definitions and recommendations. Medical literature on ultrasound vascular access was reviewed from January 1985 to October 2010. The GRADE and the GRADE-RAND methods were utilised to develop recommendations. RESULTS: The recommendations following the conference suggest the advantage of 2D vascular screening prior to cannulation and that real-time ultrasound needle guidance with an in-plane/long-axis technique optimises the probability of needle placement. Ultrasound guidance can be used not only for central venous cannulation but also in peripheral and arterial cannulation. Ultrasound can be used in order to check for immediate and life-threatening complications as well as the catheter's tip position. Educational courses and training are required to achieve competence and minimal skills when cannulation is performed with ultrasound guidance. A recommendation to create an ultrasound curriculum on vascular access is proposed. This technique allows the reduction of infectious and mechanical complications. CONCLUSIONS: These definitions and recommendations based on a critical evidence review and expert consensus are proposed to assist clinicians in ultrasound-guided vascular access and as a reference for future clinical research.

Incidence of laryngospasm and bronchospasm in pediatric adenotonsillectomy.

OBJECTIVES/HYPOTHESIS: To evaluate and describe airway complications in pediatric adenotonsillectomy. STUDY DESIGN: Retrospective case-control study. METHODS: A chart review of patients that underwent adenotonsillectomy between 2006 and 2010 was performed. Perioperative complications, patient characteristics, and surgeon and anesthesia technique were recorded. RESULTS: A total of 682 charts were reviewed. Eleven cases (1.6%) of laryngospasm were identified: one was preoperative, seven occurred in the operating room postextubation, and three occurred in the recovery area. Four patients were given succinylcholine, one was reintubated, and the other cases were managed conservatively. Mean age of patients with laryngospasm was 5.87 years (standard deviation [SD], 4.01; 1.9-15.8 years). There were 12 cases (1.8%) of bronchospasm; all were treated with nebulized albuterol. Mean age of patients with bronchospasm was 5.81 years (SD, 4.17; 1.8-14.1 years). Overall, 22 patients required antiemetics (3.3%), 19 required albuterol (2.9%), and five required racemic epinephrine (0.8%). Compared to the children without airway complications, there was no difference in age, weight, American Society of Anesthesiologists status, length of surgery, need for admission, and anesthesia technique in those that had laryngospasm. Patients with bronchospasm, compared to the patients without complications, had faster surgeries (P < .05), were more likely to have underlying asthma (P < .05), and were more likely to be admitted (P < .05). There were no unexpected admissions or other morbidities. CONCLUSIONS: The rates of laryngospasm (1.6%) and bronchospasm (1.8%) are significantly lower than reported in the literature, reflecting refinements in modern anesthesia/surgical technique. Knowledge of at-risk patients can facilitate planning to potentially reduce the incidence of perioperative airway complications during adenotonsillectomy.

Transmission of zoonoses in xenotransplantation: porcine endogenous retroviruses from an immunological and molecular point of view.

BACKGROUND AND OBJECTIVES: Pigs offer an unlimited source of xenografts for humans. The use of transplants from animal origin offers a potential solution to the limited supply of human organs and tissues. However, like many other mammalian species, pigs harbor porcine endogenous retrovirus (PERV), which are encoded in their genomic DNA and are assumed to have been integrated into the porcine germ line more than 7.6 × 106 years ago and showing that the age correlates with the time of separation between pigs and peccaries 7.4 × 106 years ago. The ability of the PERV to infect human cells in vitro has heightened safety concerns regarding the transmission of PERV to pig xenograft recipients. In this study, we detected PERV-AC recombinant virus in porcine genomic DNA that may have resulted from exogenous viral recombination. Infectious risk in xenotransplantation will be defined by the activity of PERV loci in vivo. We identified unique Haemophilus aegyptius III HaeIII gag restriction fragment length polymorphism (RFLP) profiles resulting from single nucleotide polymorphisms; these were found only in animals that produced human tropic PERV. MATERIALS AND METHODS: Porcine tissues were analyzed using validated assays specific for PERV: polymerase chain reaction (PCR) (for PERV DNA), RT-PCR (for PERV RNA), cell culture, RFLP analysis, and sequence analysis. CONCLUSIONS AND INTERPRETATION: These findings have demonstrated that the presence of both DNA and RNA forms of porcine endogenous retrovirus in porcine tissues needs to be carefully considered when the infectious disease potential of xenotransplantation is being assessed.

Geographical location and age affects the incidence of parasitic infestations in school children.

UNLABELLED: Environmental factors affect the dissemination and distribution of intestinal parasites in human communities. To comprehend the prevalence of parasitic infestation and to examine whether geographical location and age also influence the prevalence of infection, fecal samples from 195 school children (rural = 95; male = 39; female = 56) (urban = 100; male = 60; female = 40) of five age groups ranging from 5 to 11 years in two different socio-economic zones (rural and urban) were screened for specific intestinal parasites using standard histological techniques. Percentage incidences of parasitic species found in fecal wet mounts and concentrates in rural children were Entamoeba coli (25.3%), Giardia lamblia (17.9%), Blastocystis hominis (14.7%), Entamoeba histolytica (4.2%), Iodamoeba butschlii (1.1%), Hymenolepis nana (1.1%) and Ascaris lumbricoides (1.1%). Whereas the percentage incidences among urban children were E. coli (26%), A. lumbricoides (21%), B. hominis (18%), G. lamblia (14%), T. trichiura (8%), I. butschlii (4%) and A. duodenale (1%). Such findings may be related to dietary differences, living conditions and the greater use of natural anti-helminthic medicinal plants in rural communities. These results are important for both epidemiological data collection and for correlating dietary differences to intestinal parasitic diseases. AIMS: We chose to investigate whether geographical location and age affect the prevalence and distribution of intestinal parasites among school children from two separate regions (rural and urban) in areas surrounding, Chennai, Tamil Nadu, India. SETTINGS AND DESIGN: A study of the prevalence of parasitic infestations was undertaken among primary school children, in rural and urban communities around Chennai, Tamil Nadu, India. MATERIALS AND METHODS: Faecal sample collection, direct microscopic techniques, macroscopic examination and concentration techniques for identifying the parasites. STATISTICAL ANALYSIS USED: Percentage incidences of parasitic species found in faecal wet mounts and concentrates were done instead of statistical analyses. RESULTS: Both macroscopic and microscopic examinations of faecal samples revealed that the overall percentage prevalence of parasite species encountered in rural children were Entamoeba coli (25.3%), G. lamblia (17.9%), B. hominis (14.7%), Entamoeba histolytica (4.2%), I. butschlii (1.1%), H. nana (1.1%), Ascaris lumbricoides (1.1%). The prevalence among urban children were E. coli (26%), A. lumbricoides (21%), B. hominis (18%), G. lamblia (14%), T. trichiura (8%), I. butschlii (4%) and A. duodenale (1%). Overall, comparative significant differences were noted between rural and urban children for E. histolytica (4.2 vs. 14%), G. lamblia (17.9 vs. 14%), A. lumbricoides (1.1 vs. 21%) and T. trichiura (0 vs. 8%), with the major difference being the much higher occurrence of A. lumbricoides and T. trichiura infections in urban children. CONCLUSIONS: One of the greatest challenges for healthcare professionals is the prevention and treatment of protozoal and helminthic parasitic infections. From our study we conclude that the prevalence of different pathogenic species of amoeba such as Entamoeba histolytica (4.2 vs. 0%) and G. lamblia (17.9 vs. 14%), (P value was equal to 1) was significantly higher among rural children compared to children from urban areas. In contrast, the prevalence of nematodes such as A. lumbricoides (21% vs. 1.1%), T. trichiura (8% vs. 0%) and A. duodenale (1%) was also significantly higher among rural children.

Aggressive bronchoscopic management of plastic bronchitis.

Plastic bronchitis or "Bronchitis Plastica" is a rare disease characterized by the formation of thick, tenacious, arborizing mucofibrinous tracheobronchial casts that result in life-threatening airway obstruction and pulmonary failure. We review three children who developed recurrent plastic bronchitis after undergoing a Fontan procedure for single ventricle physiology. Case series of three patients with plastic bronchitis at a tertiary referral children's hospital. All patients required repeated bronchoscopies, one requiring four separate ones over a week's period, for removal of the rigid casts. Extra-corporal membrane oxygenation (ECMO) was needed in two children because of severe respiratory failure. All were also managed with adjunctive intensive medical support. Pulmonary function returned to normal in all children, but recurred 2 months later in one who subsequently expired due to pulmonary failure. Plastic bronchitis is an unusual condition of unknown cause that occurs in multiple clinical settings, but especially in those children who have undergone a Fontan operation. Management of this distressing situation is difficult and early diagnosis and aggressive measures to remove rigid casts combined with intensive medical care are necessary. The intrinsic cardiopulmonary physiology of children with Fontan procedures, including the risk of arrhythmias, hypo-oxygenation, and pulmonary hypertension make this condition even more complex.

Multidisciplinary management of expanding bilateral neck hematomas in a patient with Hemophilia A with high-titer inhibitor.

There are multiple modalities by which trauma occurs to the neck. One of these includes minor suction trauma which usually results in a superficial bruising of the skin. While this usually self-resolves, patients with hemophilia are at higher risk for the development of bleeding from such trauma. Hematomas of the head and neck in patients with hemophilia have seldom been reported. We report a unique case of expanding bilateral neck hematomas secondary to suction trauma in a patient with Hemophilia A with high-titer inhibitor and highlight the importance of a multidisciplinary approach in the management of this complex patient.

Airway responses to desflurane during maintenance of anesthesia and recovery in children with laryngeal mask airways.

BACKGROUND: We sought to characterize the airway responses to desflurane during maintenance of and emergence from anesthesia in children whose airways were supported with laryngeal mask airways (LMAs). METHODS/MATERIALS: Four hundred healthy children were randomized in a 3 : 1 ratio to either desflurane or isoflurane (reference group) during anesthetic maintenance. After induction of anesthesia, anesthesia was maintained with the designated anesthetic. The investigator chose the airway (LMA and facemask), ventilation strategy and when to remove the LMA. The incidence of airway events during maintenance, emergence and recovery was recorded. RESULTS: Ninety percent of children received LMAs. The frequency of major airway events after desflurane (9%) was similar to that after isoflurane (4%) (number needed to harm [NNH] 20), although the frequency of major events after the LMA was removed during deep desflurane anesthesia (15%) was greater than during awake removal (5%) (NNH 10) (P < 0.006) and during deep isoflurane removal (2%) (NNH 8) (P < 0.03). The frequency of airway events of any severity after desflurane was greater than that after isoflurane (39% vs 27%) (P < 0.05). The frequencies of laryngospasm and coughing of any severity after desflurane were greater than those after isoflurane (13% vs 5% and 26% vs 14%, respectively) (P < 0.05). CONCLUSIONS: When an LMA is used during desflurane anesthesia in children, fewer airway events occur when it is removed when the child is awake. Although the time to discharge from recovery was not delayed and no child required overnight admission, caution should be exercised when using an LMA in children who are anesthetized with desflurane.

Acute pain management in children.

The greatest advance in pediatric pain medicine is the recognition that untreated pain is a significant cause of morbidity and even mortality after surgical trauma. Accurate assessment of pain in different age groups and the effective treatment of postoperative pain is constantly being refined; with newer drugs being used alone or in combination with other drugs continues to be explored. Several advances in developmental neurobiology and pharmacology, knowledge of new analgesics and newer applications of old analgesics in the last two decades have helped the pediatric anesthesiologist in managing pain in children more efficiently. The latter include administering opioids via the skin and nasal mucosa and their addition into the neuraxial local anesthetics. Systemic opioids, nonsteroidal anti-inflammatory agents and regional analgesics alone or combined with additives are currently used to provide effective postoperative analgesia. These modalities are best utilized when combined as a multimodal approach to treat acute pain in the perioperative setting. The development of receptor specific drugs that can produce pain relief without the untoward side effects of respiratory depression will hasten the recovery and discharge of children after surgery. This review focuses on the overview of acute pain management in children, with an emphasis on pharmacological and regional anesthesia in achieving this goal.

Detection of porcine rotavirus from tissue and faecal specimens.

Porcine small intestinal sub-mucosa is a cell-free collagen matrix that has demonstrated its ability as a scaffold material. Transplantation poses special hazards because grafted tissues and organs transmit pathogens efficiently, especially viruses. Rotavirus is thought to be confined to the intestine, causing acute diarrhoea. The purpose of this study was to evaluate the porcine intestinal tissue scaffold for Rotavirus and to study the incidence of this virus among pig herds. Only one isolate was successfully adapted to grow in cell line MA 104 from faecal samples. This isolate was further confirmed by reverse transcriptase polymerase chain reaction and sequence analysis.

l-Ascorbic acid partially protects two cycles of cisplatin chemotherapy-induced testis damage and oligo-astheno-teratospermia in a mouse model.

Cisplatin (cis-diaminedichloroplatinum-II) is a widely used antineoplastic agent in the treatment of a variety of cancers. The aim of the present study was to investigate the testicular toxicity of cisplatin in mice at human therapeutic dose-levels, and to investigate any protective effects of concomitantly administered l-ascorbic acid (i.p.; 10 mg/kg). Adult male BALB/C mice (13-15-week-old) were treated (i.p.) with two cycles of 5 days each of cisplatin with 17 days of recovery period between cycles, as follows: Group I (G-I) - water (N = 10); Group II (G-II) -L-ascorbic acid (N = 6); Group III (G-III) - 1mg/kg (N = 6); Group IV (G-IV) - 1 mg/kg + L-ascorbic acid (N = 6); Group V (G-V) - 2.5 mg/kg (N = 6); and Group VI (G-VI) - 2.5 mg/kg + L-ascorbic acid (N = 8). All animals were sacrificed on third day after the last treatment. The testis weight was decreased in a dose-dependent pattern (G-III - 44% and G-V - 54% against G-I), but l-ascorbic acid (10 mg/kg) recovered the lost weight in G-VI up to 32% against G-V (p<0.05). Seminiferous tubular pathology was indicated by vacuoles, epithelial gaps, epithelial sloughing, delayed spermiation, malorientation of spermatids, germ cell degeneration, phagocytosis of spermatids, multinucleated germ cell formation and atrophy. Structurally abnormal tubules (G-III - 33%; G-V - 100%) were induced, and protective effects were seen in G-IV (77%) and G-VI (25%; p<0.05). The tubular diameter was decreased in G-III-VI, but recovery was seen only in G-IV. The epithelial height was decreased in G-III, G-V and G-VI and the recovery was seen only in G-VI. The sperm count was decreased up to 53% and 71% against control in G-III and G-V, respectively, and recovery up to 47% and 61% was observed in G-IV and G-VI, respectively. The sperm motility was decreased up to 56% and 63% against control in G-III and G-V, respectively, and recovery was only marginal in G-IV and G-VI (p>0.05). Total sperm abnormalities were increased in G-III-V (274%, 156% and 232%, respectively, p<0.05) and l-ascorbic acid protected the effect in G-VI up to 156% (p<0.05). In conclusion, at human therapeutic dose-levels, cisplatin induces testicular damage and spermato-toxicity. l-Ascorbic acid only partially nullifies the gonadotoxic effects of cisplatin.