In vitro longitudinal lumbar spinal cord preparations to study sensory and recurrent motor microcircuits of juvenile mice.

In vitro spinal cord preparations have been extensively used to study microcircuits involved in the control of movement. By allowing precise control of experimental conditions coupled with state-of-the-art genetics, imaging, and electrophysiological techniques, isolated spinal cords from mice have been an essential tool in detailing the identity, connectivity, and function of spinal networks. The majority of the research has arisen from in vitro spinal cords of neonatal mice, which are still undergoing important postnatal maturation. Studies from adults have been attempted in transverse slices, however, these have been quite challenging due to the poor motoneuron accessibility and viability, as well as the extensive damage to the motoneuron dendritic trees. In this work, we describe two types of coronal spinal cord preparations with either the ventral or the dorsal horn ablated, obtained from mice of different postnatal ages, spanning from preweaned to 1 mo old. These semi-intact preparations allow recordings of sensory-afferent and motor-efferent responses from lumbar motoneurons using whole cell patch-clamp electrophysiology. We provide details of the slicing procedure and discuss the feasibility of whole cell recordings. The in vitro dorsal and ventral horn-ablated spinal cord preparations described here are a useful tool to study spinal motor circuits in young mice that have reached the adult stages of locomotor development.NEW & NOTEWORTHY In the past 20 years, most of the research into the mammalian spinal circuitry has been limited to in vitro preparations from embryonic and neonatal mice. We describe two in vitro longitudinal lumbar spinal cord preparations from juvenile mice that allow the study of motoneuron properties and respective afferent or efferent spinal circuits through whole cell patch clamp. These preparations will be useful to those interested in the study of microcircuits at mature stages of motor development.

Effect of aging on H-reflex response to fatigue.

Injury as a result of tripping is relatively common among older people. The risk of falling increases with fatigue and of importance is the ability to dorsiflex the foot through timely activation of the tibialis anterior (TA) muscle to ensure the foot clears the ground, or an obstacle, during the swing phase of walking. We, therefore, questioned whether the muscle spindle input to the motoneurons alters with ongoing fatigue in older people. We electrically stimulated the common peroneal nerve to assess the TA primary afferent efficacy using H-reflex before, immediately following and after a fatiguing maximal isometric contraction. M-response was kept unchanged throughout the experiment to ensure a similar stimulus intensity was delivered across time points. H-reflex increased significantly while the TA muscle was in a state of fatigue for the younger participants but tended to decrease with increasing age. The main contributor to the tonicity of TA muscle, i.e., excitatory synapses of spindle primary endings of motoneurons that innervate TA muscle, tend to lose their efficacy during fatigue in the older individuals but increased efficiency in the majority of the younger people. Since TA muscle is the main dorsiflexor of the foot and it needs to be active during the swing phase of stepping to prevent tripping, older individuals become more susceptible to falling when their muscles are fatigued. This finding may help improve devices/treatments to overcome the problem of tripping among older individuals.

Exploring the receptor origin of vibration-induced reflexes.

STUDY DESIGN: An experimental design. OBJECTIVES: The aim of this study was to determine the latencies of vibration-induced reflexes in individuals with and without spinal cord injury (SCI), and to compare these latencies to identify differences in reflex circuitries. SETTING: A tertiary rehabilitation center in Istanbul. METHODS: Seventeen individuals with chronic SCI (SCI group) and 23 participants without SCI (Control group) were included in this study. Latency of tonic vibration reflex (TVR) and whole-body vibration-induced muscular reflex (WBV-IMR) of the left soleus muscle was tested for estimating the reflex origins. The local tendon vibration was applied at six different vibration frequencies (50, 85, 140, 185, 235, and 265 Hz), each lasting for 15 s with 3-s rest intervals. The WBV was applied at six different vibration frequencies (35, 37, 39, 41, 43, and 45 Hz), each lasting for 15 s with 3-s rest intervals. RESULTS: Mean (SD) TVR latency was 39.7 (5.3) ms in the SCI group and 35.9 (2.7) ms in the Control group with a mean (95% CI) difference of -3.8 (-6.7 to -0.9) ms. Mean (SD) WBV-IMR latency was 45.8 (7.4) ms in the SCI group and 43.3 (3.0) ms in the Control group with a mean (95% CI) difference of -2.5 (-6.5 to 1.4) ms. There were significant differences between TVR latency and WBV-IMR latency in both the groups (mean (95% CI) difference; -6.2 (-9.3 to -3.0) ms, p = 0.0001 for the SCI group and -7.4 (-9.3 to -5.6) ms, p = 0.011 for Control group). CONCLUSIONS: The results suggest that the receptor of origin of TVR and WBV-IMR may be different.

Motor units as tools to evaluate profile of human Renshaw inhibition.

KEY POINTS: To uncover the synaptic profile of Renshaw inhibition on motoneurons, we stimulated thick motor axons and recorded from voluntarily-activated motor units. Stimuli generated a direct motor response on the whole muscle and an inhibitory response in active motor units. We have estimated the profile of Renshaw inhibition indirectly using the response of motor unit discharge rates to the stimulus. We have put forward a method of extrapolation that may be used to determine genuine synaptic potentials as they develop on motoneurons. These optimized techniques can be used in research and in clinics to fully appreciate Renshaw cell function in various neurological disorders. ABSTRACT: Although Renshaw inhibition (RI) has been extensively studied for decades, its precise role in motor control is yet to be discovered. One of the main handicaps is a lack of reliable methods for studying RI in conscious human subjects. We stimulated the lowest electrical threshold motor axons (thickest axons) in the tibial nerve and analysed the stimulus-correlated changes in discharge of voluntarily recruited low-threshold single motor units (SMUs) from the soleus muscle. In total, 54 distinct SMUs from 12 subjects were analysed. Stimuli that generated only the direct motor response (M-only) on surface electromyography induced an inhibitory response in the low-threshold SMUs. Because the properties of RI had to be estimated indirectly using the background discharge rate of SMUs, its profile varied with the discharge rate of the SMU. The duration of RI was found to be inversely proportional to the discharge rate of SMUs. Using this important finding, we have developed a method of extrapolation for estimating RI as it develops on motoneurons in the spinal cord. The frequency methods indicated that the duration of RI was between 30 and 40 ms depending on the background firing rate of the units, and the extrapolation indicated that RI on silent motoneurons was ∼55 ms. The present study establishes a novel methodology for studying RI in human subjects and hence may serve as a tool for improving our understanding of the involvement of RI in human motor control.

Medium latency excitatory reflex of soleus re-examined.

We aimed to study the receptor origin and postsynaptic potential profile of the medium latency reflex (MLR) response that develops in the soleus muscle when common peroneal nerve of antagonist tibialis anterior (TA) muscle is electrically stimulated. To achieve this aim, we electrically stimulated common peroneal nerve and recorded surface electromyography (SEMG) responses of soleus and TA muscles of informed volunteers. Additionally, we recorded intramuscular EMG from the soleus muscle. Stimulation of common peroneal nerve induced a direct motor response (M-response) in the TA and MLR in SEMG of the soleus. Using voluntarily-activated single motor units (SMUs) from the soleus muscle we noted that there were two distinct responses following the stimulus. The first response was a reciprocal inhibitory reflex probably originating from the antagonist muscle spindle primary (Ia) afferents. This was followed by an indirect reflex response activated by the contraction of the TA muscle during the M-response. This contraction generated a rapid acceleration in the direction of dorsiflexion hence inducing a stretch stimulus on soleus muscle. The response of soleus to this stimulus was a stretch reflex. We suggest that this stretch reflex is the main contributor to the so-called soleus MLR in the literature. This study illustrated the importance of using SMUs and also using discharge-rate based analysis for closely examining previously 'established' reflexes.

A new colorimetric method for rapid detection of ethambutol and streptomycin resistance in Mycobacterium tuberculosis: crystal violet decolorization assay (CVDA).

Streptomycin (STR) and ethambutol (EMB) are important drugs used for the treatment of tuberculosis. There is a need for fast, reliable and inexpensive methods for detecting resistance to these drugs. The aim of this study was to evaluate the performance of the crystal violet decolorization assay (CVDA) for the detection of STR and EMB resistance that is important drugs in tuberculosis treatment. In this study, drug susceptibility testing was performed on 140 Mycobacterium tuberculosis isolates provided from nine centers. Three tubes were used for each isolate. One of the tubes had a concentration of 2 mg/L STR and the other 5 mg/L EMB. The third was drug-free control tube. Sensitivity, specificity, positive predictive value (PPD), negative predictive value (NPD) and agreement for STR were found to be 81.8%, 94.6%, 87.8%, 91.5% and 90.57%, respectively. For EMB, sensitivity, specificity, PPD, NPD, and agreement were found to be 76%, 98.23%, 90.47%, 94.87% and 94.2%, respectively. The results were obtained in 11.3 ± 2.7 days (8-21 days). CVDA is rapid, reliable, inexpensive, and easy to perform for rapid detection of STR and EMB resistance, and it could be adapted for drug susceptibility testing.

Optimal location for eliciting the tibial H-reflex and motor response.

INTRODUCTION: Although there are numerous protocols to adjust the amplitude of the Hoffmann reflex (H-reflex) relative to the size of the direct motor response (M-response), the optimal stimulating location has not been described. We sought to determine the optimal positioning of the stimulating cathode when evoking the tibial nerve H-reflex and M-response. METHODS: A small cathode was placed on defined points in the popliteal fossa while an anode was fixed on the patella. The tibial nerve was stimulated electrically, and the response of the soleus muscle was recorded using intramuscular and surface electromyography. RESULTS: We found that more-lateral points along a line drawn across the popliteal fossa were the best locations to obtain only the M-response, whereas stimulating the midpoint was optimal for obtaining only the H-reflex. DISCUSSION: By using specified locations for electrical stimulation to evoke H-reflex and M-response, the functionality of the tibial nerve can be assessed. Muscle Nerve 58:828-833, 2018.

Predictors of fatality in pandemic influenza A (H1N1) virus infection among adults.

BACKGROUND: The fatality attributed to pandemic influenza A H1N1 was not clear in the literature. We described the predictors for fatality related to pandemic influenza A H1N1 infection among hospitalized adult patients. METHODS: This is a multicenter study performed during the pandemic influenza A H1N1 [A(H1N1)pdm09] outbreak which occurred in 2009 and 2010. Analysis was performed among laboratory confirmed patients. Multivariate analysis was performed for the predictors of fatality. RESULTS: In the second wave of the pandemic, 848 adult patients were hospitalized because of suspected influenza, 45 out of 848 (5.3%) died, with 75% of fatalities occurring within the first 2 weeks of hospitalization. Among the 241 laboratory confirmed A(H1N1)pdm09 patients, the case fatality rate was 9%. In a multivariate logistic regression model that was performed for the fatalities within 14 days after admission, early use of neuraminidase inhibitors was found to be protective (Odds ratio: 0.17, confidence interval: 0.03-0.77, p=0.022), nosocomial infections (OR: 5.7, CI: 1.84-18, p=0.013), presence of malignant disease (OR: 3.8, CI: 0.66-22.01, p=0.133) significantly increased the likelihood of fatality. CONCLUSIONS: Early detection of the infection, allowing opportunity for the early use of neuraminidase inhibitors, was found to be important for prevention of fatality. Nosocomial bacterial infections and underlying malignant diseases increased the rate of fatality.

Antimicrobial susceptibility and resistance mechanisms of methicillin resistant Staphylococcus aureus isolated from 12 Hospitals in Turkey.

INTRODUCTION: Methicillin-resistant Staphylococcus aureus (MRSA) is one of the most important nosocomial pathogens and is also emerging in Turkish hospitals. The aim of this study was to determine the antimicrobial susceptibility profiles of MRSA isolated from Turkish hospitals. MATERIALS AND METHODS: A total of 397 MRSA strains isolated from 12 hospitals in Turkey were included to present study. Antimicrobial susceptibilities were tested using agar dilution method. Presence of ermA, ermB, ermC, msrA, tetM, tetK, linA and aac-aph genes were studied by PCR. RESULTS: All strains were susceptible to vancomycin and linezolid. The susceptibility rates for fusidic acid, lincomycin, erythromycin, tetracyclin, gentamycin, kanamycin, and, ciprofloxacin were 91.9%, 41.1%, 27.2%, 11.8%, 8.5%, 8.3% and 6.8%, respectively. Lincomycin inactivation was positive for 3 isolates. Of 225 erythromycin resistant isolates 48 had ermA, 20 had ermC, and 128 had ermA-C. PCR was negative for 15 strains. Of 3 isolates with lincomycin inactivation one had linA and msrA. Of 358 gentamycin resistant isolates 334 had aac-aph and 24 were negatives. Among 350 tetracyclin resistant isolates 314 had tetM. Of 36 tetM negative isolates 10 had tetK. CONCLUSION: MRSA isolates from Turkish hospitals were multiresistant to antimicrobials. Quinolone and gentamycin resistance levels were high and macrolide and lincosamide resistance were relatively low. Susceptibility rates for fusidic asid were high. Linezolide and vancomycin resistance are not emerged. The most common resistance genes were ermA, tetM and aac-aph. Evolution of antimicrobial susceptibilities and resistance genes profiles of MRSA isolates should be surveyed at regional and national level for accurate treatment of patients and to control dissemination of resistance genes.

[Heterogeneous macrolide resistance in methicillin-resistant Staphylococcus aureus isolates: investigation of resistance mechanisms and clonality]. / Metisiline dirençli Staphylococcus aureus izolatlarinda heterojen makrolid direnci: Direnç mekanizmasinin ve klonalitenin arastirilmasi

Staphylococcus aureus is one of the most important pathogens leading to hospital infections and has ability to gain multiple resistance to most of the antibiotics used as well as ability to spread clonally. In this study we aimed to investigate the mechanism of heterogenous macrolide resistance and clonality of 41 methicillin-resistant S.aureus (MRSA) strains isolated from clinical samples of inpatients between 2006 and 2008 in a tertiary care training hospital. Heterogenous macrolide resistance is defined as the isolates with colonies in the inhibiton zone of erythromycin, azithromycin, claritromycin discs, and named as hMLSB (heterogeneous macrolides, lincosamides, streptogramin-B) phenotype. A total of 63 macrolide-resistant MRSA strains isolated during the same period with non-hMLSB phenotype were included in the study as a control group. The susceptibilties of isolates to methicillin, erythromycin, azithromycin, claritromycin, clindamycin, quinupristin-dalfopristin, penicillin, vancomycin, teicoplanin, linezolide, gentamicin, amikacin, ciprofloxacin, trimethoprim-sulphamethoxazole, chloramphenicol, rifampin, tetracycline and telithromycin were determined by disc diffusion method according to the CLSI criteria. hMLSB isolates were also tested for erythromycin, clindamycin and quinupristin-dalfopristin susceptibility by Vitek-2 automated system (bioMerieux, France). Presence of erm(A), erm(B), erm(C), msr(A/B) resistance genes were investigated by PCR. The regulatory region of the erm(A) gene, which was found to be the only molecular mechanism of hMLSB, was sequenced. Sequence analysis of regulatory regions showed deletion of "guanine" base at position 149 in 34 strains and two point mutations namely "C368G" and "T377C" in 16 strains. Pulsed-field gel electrophoresis (PFGE) analysis indicated presence of a common clone and its variants; however this clone was also common among control group strains. Interestingly all heterogeneous macrolide resistant isolates were reported as susceptible to erythromycin, clindamycin, quinupristin-dalfopristin by the automated system. As a result a clone specific to heterogenous macrolide resistant strains was not detected. Although all hMLSB strains had erm(A) gene, a specific and common genetic modification could not be found in regulatory region of the isolates. The most important finding of this study is the detection of inability of the automated system to properly reflect the hMLSB phenotype. In addition it was suggested that longer incubation (at least 24 hours) of the antibiotic susceptibility test plates could help identification of hMLSB phenotype in disc diffusion tests.

[Investigation of extensive drug resistance in multidrug resistance tuberculosis isolates]. / Çok Ilaca Dirençli Tüberküloz Izolatlarinda Yaygin Ilaç Direncinin Arastirilmasi

Increasing number of drug resistant tuberculosis (TB) cases, observed in recent years, is an important public health problem. Extensively drug resistant TB (XDR-TB) is the development of resistance against any fluoroquinolones and at least one of the injectable second line anti-TB drugs in addition to resistance against isoniazide and rifampicin which are the first line anti-TB drugs [definition of multidrug resistant TB (MDR-TB)]. Anti-TB therapy failed with first-line anti-TB drugs due to MDR-TB cases is being planned according to second-line anti-TB drug susceptibility test results if available and if not, standart treatment protocols are used. Although it is recommended that individual anti-TB therapy should be designed according to the isolate's susceptibility test results, standart therapeutic protocols are always needed since second-line anti-TB drug susceptibility testing generally could not be performed in developing countries like Turkey. For this reason, nationwide and regional surveillance studies to determine the resistance patterns are always needed to make decisions about the standard therapy algorithms. In this study, it was aimed to investigate the presence of extensive drug resistance among 81 MDR-TB isolates obtained from various health care facilities from Istanbul, Izmir and Manisa and to determine the XDR-TB incidence in Marmara and Aegean regions. Furthermore, we aimed to provide epidemiological data to clinicians to support their choice of second-line anti-TB drugs for MDR-TB infections. Susceptibility testing of isolates for the first and the second-line anti-TB drugs were performed by using modified Middlebrook 7H9 broth in fluorometric BACTEC MGIT 960 system (Becton Dickinson, USA). Eighty-one MDR-TB isolates included in this study were isolated from 43 (53.1%) patients residing in Istanbul, 26 (32.1%) in Izmir and 12 (14.8%) in Manisa provinces. We could not find any isolate consistent with XDR-TB definition in this study. Second-line drug resistance rates of MDR-TB isolates to amikacin and kanamycin were 1.2%, ofloxacin and levofloxacin were 2.5%, capreomycin was 14.8%, ethionamide was 37% whereas linezolid resistance was not detected. Statistically significant correlation was detected between resistance rates of these antibiotic pairs; levofloxacin-ofloxacin (p< 0.01), amikacin-kanamycin (p= 0.01) and streptomycin-ethionamide (p= 0.04). In our study, extensive drug resistance was not encountered in any MDR-TB isolates while high resistance rates was observed against ethionamide and capreomycin. It can be concluded that parenteral aminoglycosides amikasin and kanamycin, fluoroquinolones and linezolid seemed to be reliable anti-TB agents in MDR-TB treatment, however, further larger scale studies are needed.

[t030 is the most common spa type among methicillin-resistant Staphylococcus aureus strains isolated from Turkish hospitals]. / t030, Türkiye'deki Hastanelerden Izole Edilen Metisiline Dirençli Staphylococcus aureus Izolatlari Arasinda En Yaygin spa Tipidir.

Staphylococcus aureus is one of the most frequent agents causing hospital infections. S.aureus has a great ability to adapt itself to variety of conditions and successful clones can be epidemic and even pandemic by its ability spread from one continent to another. The aims of this study were to detect spa types of 397 methicillin-resistant S.aureus (MRSA) strains isolated from 12 centers in different geographical regions of Turkey from 2006 to 2008, and to investigate their clonality by PFGE and MLST typing. Additionally, 91 MRSA from four of those 12 centers isolated during 2011 were also studied for their spa types. PFGE profiles indicated the presence of a major pulsotype, namely pulsotype A with a rate of 91.4% (363/397), followed by pulsotype B (n= 18, 4.5%) and pulsotype C (n= 11, 2.8%). Among isolates tested 363 (91.4%) were SCCmec type III, 30 (7.6%) were SCCmec type IV. Sequence analysis of representative isolates revealed that ST239 (85.1%) was the most common MLST type followed by two MLST types ST737 (4%), and ST97 (2.8%), both SCCmec type IV. Two isolates were ST80 with SCCmec type IV. Of 397 isolates, 338 (85.1%) were t030, followed by t005 (2.5%) and t632 (2%). Among MRSA isolated during 2011, 64 (70.3%) of 91 were t030, 4 (4.4%) were t005, 2 (2.2%) were t015, and 2 (2.2%) were t1094. Among centers the t030 prevalence of 2006-2008 isolates ranged from 59-100%. The highest t030 prevalence was found in Ankara (100%) and lowest in Trabzon (59%) provinces which are located at central and northestern Anatolia, respectively. In Istanbul province, the prevalence of t030 was 94.5% among 2006-2008 isolates which decreased to 55.5% among 2011 isolates. Also a decrease in t030 rates was observed among samples from Konya and Trabzon but not from Aydin. Our results showed that the most common MRSA clone in Turkey is ST 239-SCCmec type III, t030 which persisted during the six years of the study period. Presence of PVL toxin gene was tested by PCR and 5 (3%) isolates found to be positive, of them two were SCCmec Type IV-ST80 and three were SCCmec Type III-ST239. This study is the largest epidemiological survey ever done in Turkey which showed presence of a hospital Turkish clone TR09 (ST239-SCCmecIII-t030) and a community clone TR10 (ST737-SCCmecIV-t005) largely disseminated in Turkey.

Effects of infliximab on bacterial translocation in experimental acute necrotizing pancreatitis.

BACKGROUND & OBJECTIVES: Translocation of bacteria from the gut is an important factor in the development of septic complications and mortality in acute pancreatitis (AP). The present study was designed to assess the effects of infliximab treatment on bacterial translocation (BT) in experimental acute necrotizing pancreatitis. METHODS: Male Sprague-Dawley rats (n=45) were allocated into three groups. AP was induced in group II (positive control, n=15) and group III (Infliximab; n=15) by retrograde injection of taurocholate into the common biliopancreatic duct. Group I rats (Sham; n=15) received normal saline infusion into the common biliopancreatic duct as placebo. Groups I and II were treated by normal saline and group III was treated with infliximab intraperitoneally on 6, 30 and 54 h after induction of pancreatitis. All surviving animals were killed 60 h after the induction of pancreatitis, and specimens were collected for amylase measurement as well as histopathologic and microbiologic examinations. RESULTS: Oedema, acinar cell necrosis, inflammatory infiltration, haemorrhage, fat necrosis and perivascular inflammation in group III rats were decreased with infliximab treatment when compared with group II (P<0.001). BT to mesentery lymph node in groups I, II and III were 20, 100 and 46 per cent, respectively. BT to peritoneum and pancreas in group III was lower than group II (P<0.05). INTERPRETATION & CONCLUSIONS: Infliximab administration resulted in beneficial effects on BT and histopathologic changes in the experimental necrotizing pancreatitis. Whether anti-TNF therapy has a role in prevention of complications of ANP needs to be established.

Pulmonary tuberculosis and tuberculous arthritis of knee joint associated with rheumatoid arthritis treated with anti-tumor necrosis factor (TNF)-alpha medication: a case report.

Tuberculosis infection (TB) is one of the most important problems for the rheumatoid arthritis (RA) patients treated with anti-TNF agents. Pulmonary tuberculosis is the most common clinic form of the TB in these patients. However, tuberculosis arthritis is very rare. We present here a 72-year-old Caucasian woman with seropositive RA, treated with etanercept/adalimumab for the last 2 years, who presented with resistant knee pain and joint effusion. We believe that this treatment caused the tuberculosis in this patient, which is the most worried complication. Interestingly, tuberculosis was in the knee joint at this time.

[Use of molecular techniques in the diagnosis of pulmonary and extrapulmonary tuberculosis]. / Akciger ve Akciger Disi Tüberküloz Tanisinda Moleküler Yöntemlerin Kullanimi

Tuberculosis, is a life-threatening infectious disease and one of the leading causes of mortality worldwide. It is a multisystemic disease, and involvement of lungs and pleura is seen in majority of patients. The global control of tuberculosis is impeded by the relatively low sensitive conventional diagnostic assays, especially for drug resistant strains. The current gold standard for tuberculosis diagnosis is the combination of culture and clinical diagnosis. Since conventional diagnostic assays require long time, are labor intensive and insufficient for species-level identification, new solutions for problems in routine diagnostic applications for tuberculosis are required. Implementation of reliable, highly specific and sensitive assays which were standardized for rapid diagnosis of Mycobacterium tuberculosis, is recommended by Centers for Disease Control and Prevention (CDC). Implementation of nucleic-acid amplification based tests (NATs) to support rapid diagnosis especially in reference laboratories or health-care clinical laboratories, which use the diagnostic algorithms denoted in CDC guidelines, meet American Thoracic Society class 2 and 3 standards and have smear positive sample counts ≥ 500 per year, is accepted as a logical and cost-effective approach. Analysis of cost-effectiveness for NATs are also required for global TB management plans. All current meta-analysis on NAT for tuberculosis indicate that in-house assays display considerable variations in sensitivity and specificity and exhibit discrepancies with commercial NATs. Evaluations also notified that the specificity for NAT is usually higher than expected in laboratory diagnosis of both forms of tuberculosis, whereas sensitivity is very low with considerable inter-assay variations. In conclusion, it is thought that current NATs are not reliable as single assays for rapid tuberculosis diagnosis for routine testing and should not be used in screening, but they are valuable methods in supporting conventional assays for clinical follow-up of patients. This review article discusses the diagnostic value of molecular methods in the evaluation of pulmonary and extrapulmonary tuberculosis in the light of the current literature.

[Investigation of a nosocomial outbreak caused by ESBL positive Klebsiella pneumoniae in neonatal intensive care unit by AP-PCR]. / Yenidogan Yogun Bakim Ünitesinde Ortaya Çikan GSBL Pozitif Klebsiella pneumoniae Nedenli Nozokomiyal Bir Salginin AP-PCR ile Arastirilmasi

In the present study, we describe an outbreak caused by extended-spectrum beta-lactamase (ESBL) producing Klebsiella pneumoniae in the neonatal intensive care unit (NICU) of a tertiary-care hospital. Nosocomial blood-stream infections were detected in three patients hospitalized in NICU. Two of the cases were transferred to NICU due to premature birth and the other due to the presence of cleft palate and retrognathia. K.pneumoniae was isolated on the 5th day of hospitalization of the first patient from umbilical swab and blood culture; on the 15th day of hospitalization of the second patient from blood culture and on the 7th day of hospitalization of the third patient from blood culture. The isolates were identified by automated API Rapid ID 32 Staph (BioMerieux, France) system in addition to conventional laboratory methods. Antibiotic susceptibilities of the isolates were determined by using Kirby-Bauer disk diffusion method according to Clinical and Laboratory Standards Institute (CLSI) criteria. The same antibiotic susceptibility patterns were detected in all isolates. Active surveillance cultures included environmental sampling from surfaces of NICU, laryngoscopes, ventilators and connections of ventilators, stethoscopes, nebulizers, aspiration tubings, disinfectant solutions, couveuse and couveuse distilled water. Two ESBL producing K.pneumoniae strains, presenting the same antibiotic susceptibility pattern with the clinical strains, were isolated from one couveuse distilled water sample and one aspiration tubing. All of the K.pneumoniae isolates were resistant to amoxycillin-clavulonic acid, cefazolin, cefepime, ceftriaxone, ceftazidime, cefuroxime, aztreonam and trimetoprim-sulphametoxazole and susceptible to cefoxitin, imipenem, meropenem, gentamicin, tobramycin, amikacin, netilmisin, tetracycline, ciprofloxacin and chloramphenicol. Arbitrarily primed polymerase chain reaction (AP-PCR) analysis done with M13 primer revealed the same genotype for the patient and environmental K.pneumoniae isolates. It was concluded that AP-PCR which is a simple, rapid and cheap method for the determination of genetic relatedness between isolates, can be applied for the detailed evaluation of nosocomial outbreaks to detect the source of infection and control the dissemination of the outbreak.

The Effects of Optogenetic Activation of Astrocytes on Spike-and-Wave Discharges in Genetic Absence Epileptic Rats.

Background: Absence seizures (petit mal seizures) are characterized by a brief loss of consciousness without loss of postural tone. The disease is diagnosed by an electroencephalogram (EEG) showing spike-wave discharges (SWD) caused by hypersynchronous thalamocortical (TC) oscillations. There has been an explosion of research highlighting the role of astrocytes in supporting and modulating neuronal activity. Despite established in vitro evidence, astrocytes' influence on the TC network remains to be elucidated in vivo in the absence epilepsy (AE). Purpose: In this study, we investigated the role of astrocytes in the generation and modulation of SWDs. We hypothesize that disturbances in astrocytes' function may affect the pathomechanism of AE. Methods: To direct the expression of channelrhodopsin-2 (ChR2) rAAV8-GFAP-ChR2(H134R)-EYFP or to control the effect of surgical intervention, AAV-CaMKIIa-EYFP was injected into the ventrobasal nucleus (VB) of the thalamus of 18 animals. After four weeks following the injection, rats were stimulated using blue light (~473 nm) and, simultaneously, the electrophysiological activity of the frontal cortical neurons was recorded for three consecutive days. The animals were then perfused, and the brain tissue was analyzed by confocal microscopy. Results: A significant increase in the duration of SWD without affecting the number of SWD in genetic absence epileptic rats from Strasbourg (GAERS) compared to control injections was observed. The duration of the SWD was increased from 12.50 ± 4.41 s to 17.44 ± 6.07 following optogenetic stimulation in GAERS. The excitation of the astrocytes in Wistar Albino Glaxo Rijswijk (WAG-Rij) did not change the duration of SWD; however, stimulation resulted in a significant increase in the number of SWD from 18.52 ± 11.46 bursts/30 min to 30.17 ± 18.43 bursts/30 min. Whereas in control injection, the duration and the number of SWDs were similar at pre- and poststimulus. Both the background and poststimulus average firing rates of the SWD in WAG-Rij were significantly higher than the firing recorded in GAERS. Conclusion: These findings suggest that VB astrocytes play a role in modulating the SWD generation in both rat models with distinct mechanisms and can present an essential target for the possible therapeutic approach for AE.

Spinal premotor interneurons controlling antagonistic muscles are spatially intermingled.

Elaborate behaviours are produced by tightly controlled flexor-extensor motor neuron activation patterns. Motor neurons are regulated by a network of interneurons within the spinal cord, but the computational processes involved in motor control are not fully understood. The neuroanatomical arrangement of motor and premotor neurons into topographic patterns related to their controlled muscles is thought to facilitate how information is processed by spinal circuits. Rabies retrograde monosynaptic tracing has been used to label premotor interneurons innervating specific motor neuron pools, with previous studies reporting topographic mediolateral positional biases in flexor and extensor premotor interneurons. To more precisely define how premotor interneurons contacting specific motor pools are organized, we used multiple complementary viral-tracing approaches in mice to minimize systematic biases associated with each method. Contrary to expectations, we found that premotor interneurons contacting motor pools controlling flexion and extension of the ankle are highly intermingled rather than segregated into specific domains like motor neurons. Thus, premotor spinal neurons controlling different muscles process motor instructions in the absence of clear spatial patterns among the flexor-extensor circuit components.

The spinal cord contains circuits of nerve cells that control how the body moves. Within these networks are interneurons that project to motor neurons, which innervate different types of muscle to contract: flexors (such as the biceps), which bend, or 'flex', the body's joints, and extensors (such as the triceps), which lead to joint extension. These motor signals must be carefully coordinated to allow precise and stable control of the body's movements. Previous studies suggest that where interneurons are placed in the spinal cord depends on whether they activate the motor neurons responsible for flexion or extension. To test if these findings were reproducible, Ronzano, Skarlatou, Barriga, Bannatyne, Bhumbra et al. studied interneurons which flex and extend the ankle joint in mice. In collaboration with several laboratories, the team used a combination of techniques to trace how interneurons and motor neurons were connected in the mouse spinal cord. This revealed that regardless of the method used or the laboratory in which the experiments were performed, the distribution of interneurons associated with flexion and extension overlapped one another. This finding contradicts previously published results and suggests that interneurons in the spinal cord are not segregated based on their outputs. Instead, they may be positioned based on the signals they receive, similar to motor neurons. Understanding where interneurons in the spinal cord are placed will provide new insights on how movement is controlled and how it is impacted by injuries and disease. In the future, this knowledge could benefit work on how neural circuits in the spinal cord are formed and how they can be regenerated.

Co-expression of plasmid-mediated quinolone resistance-qnrA1 and blaVEB-1 gene in a Providencia stuartii strain.

An extended-spectrum B-lactamase (ESBL)-producing Providencia stuartii isolate was studied. A qnrA1 gene co-expressing blaVEB-1 gene was detected. Both genes were transferred to the recipient strain. The ciprofloxacin MIC of recipient strain increased tenfold. The blaVEB-1 gene persisted in microorganisms in Turkey but it also spread with PMQR genes to other species. The combination of PMQR with multidrug resistant isolates producing ESBLs may compromise the use of valuable antibiotics. Serious efforts are necessary to detect PMQR determinants not only with common B-lactamases in widespread pathogens but also with uncommon forms that are encountered infrequently.

[Phenotypic and genotypic identification of Candida strains isolated as nosocomial pathogens]. / Hastane enfeksiyonu etkeni olarak izole edilen Candida suslarinin genotipik ve fenotipik olarak tanimlanmasi.

Over the last decade, there have been important changes in the epidemiology of Candida infections and antifungal agents used to treat these infections. In recent years, Candida species have emerged as important causes of invasive infections among patients in intensive care units. One of the main goals of this study was to evaluate the molecular epidemiology of infectious Candida species isolated in our hospital and accordingly supply data for hospital infection (HI) control. The other aim of this study was to evaluate effectiveness and practical applicability of traditional and molecular methods used to identify Candida isolates to the species level. A total of 77 Candida strains that were isolated from various clinical specimens of 60 hospitalized patients (29 male, 24 female; 7 were children) were included in the study. Fifty-seven (74%) of those isolates were defined as HI agents according to Centers for Disease Control and Prevention (CDC) criteria. The most common Candida species identified as agents of HI were C.albicans (22; 38.6%), followed by C.tropicalis (14; 24.6%), C.parapsilosis (13; 22.8%), C.glabrata (7; 12.3%) and Candida spp. (1; 1.75%). It was determined that bloodstream (26; 45.6%) and urinary tract infections (24; 42.1%) were the most frequently encountered nosocomial infections caused by Candida species. In addition it was detected that the most frequent causative agent of bloodstream infections was C.parapsilosis (10; 38.5%) and of urinary tract infections was C.albicans (12; 50%). The evaluation of advantages and disadvantages of traditional phenotypic methods [germ tube formation, chlamydospore formation in corn meal agar, growth at 45°C, colony characteristics on CHROMagar Candida medium, carbohydrate assimilation properties detected by API ID 32C (BioMerieux, France) system] and some molecular techniques [polymerase chain reaction (PCR) by using ITS-1, ITS-3 and ITS 4 primers, PCR-Restriction fragment length polymorphism (RFLP), PCRRFLP in which ITS1-ITS4 products cut by Msp I ve Bln I restriction enzymes] for the identification of Candida species revealed that CHROMagar Candida medium combined with API ID 32C kit yielded the same results (100% compatible) as molecular techniques for the species identification of Candida isolates. Since these phenotypic methods were simple and cost effective when compared to molecular techniques, they should be considered in the identification of Candida species.