Molecular characterization of multidrug-resistant Klebsiella pneumoniaeand Escherichia coliharbouring extended spectrum beta-lactamases andcarbapenemases genes at a tertiary hospital, Kenya

Background: Multidrug-resistant (MDR) Gram negative rods are increasingly being reported in sub-Saharan Africa. Molecular investigations play an important role, alongside other measures, in controlling nosocomial infections attributed to these organisms. This study aimed to determine the common extended spectrum beta-lactamases (ESBL) and carbapenemases genes, and clonal relationship in MDR Klebsiella pneumoniae and Escherichia coli. Methods: Fifty-four MDR isolates collected at the Aga Khan University hospital, Nairobi in the month of August 2012 formed the study. These were picked after an increase in the number of resistant strains during the said period was experienced. Results: blaCTXM was present in 41 (74%) of the isolates, while blaSHV was detected in 18 (33%) and blaTEM in 13 (24%) of the isolates. Nine (16.7%) of the isolates harboured all three ESBL genes and 8 (14.8%) harboured two. Eight of the isolates (all E. coli) had none of the ESBL genes tested. Two isolates harboured carbapenemases genotypes: one had blaNDM-1 and the other blaSPM. Sequencing matched CTXM-15 and TEM-1 genes in all the isolates harbouring blaCTXM and blaTEM respectively. However, there was diversity in blaSHV with SHV-11 and SHV-12 genes predominant. The isolates were non-clonal. Conclusions: The isolates mostly harboured blaCTX-M-15 while only a few had carbapenemases genes. Lack of clonality suggests these were the stable circulating strains at the time of the study

Genetic basis of rifampicin resistance in methicillin-resistant Staphylococcus aureus suggests clonal expansion in hospitals in Cape Town, South Africa

<p>Abstract</p> <p>Background</p> <p>Since 2001, several studies have reported high rifampicin resistance rates (45 - 100%) among methicillin-resistant <it>Staphylococcus aureus </it>(MRSA) isolates from South Africa. The authors previously characterised 100 MRSA isolates from hospitals in Cape Town, South Africa; forty-five percent of these isolates were rifampicin-resistant. The majority (44/45) corresponded to ST612-MRSA-IV, which is prevalent in South Africa, but has not been reported frequently elsewhere. The remaining rifampicin-resistant isolate corresponded to ST5-MRSA-I. The aim of this study was to investigate further the prevalence and genetic basis of rifampicin-resistance in MRSA isolates from hospitals in Cape Town.</p> <p>Results</p> <p>Between July 2007 and June 2011, the prevalence of rifampicin-resistant MRSA in hospitals in Cape Town ranged from 39.7% to 46.4%. Based on the results of the aforementioned study, nine ST612-MRSA-IV isolates, the rifampicin-resistant ST5-MRSA-I isolate, and two rifampicin-susceptible MRSA isolates were investigated. Four previously described ST612-MRSA-IV isolates, including two each from South Africa and Australia, were also included.</p> <p>The ST5-MRSA-I isolate carried a single mutational change, H₄₈₁Y, commonly associated with high-level rifampicin resistance. All ST612-MRSA-IV isolates carried an uncommon double amino acid substitution in RpoB, H₄₈₁N, I₅₂₇M, whilst one of the Australian ST612-MRSA-IV isolates carried an additional mutation within <it>rpoB</it>, representing a novel <it>rpoB </it>genotype: H₄₈₁N, I₅₂₇M, K₅₇₉R. All ST612-MRSA-IV isolates also shared a unique silent single nucleotide polymorphism (SNP) within <it>rpoB</it>.</p> <p>Conclusions</p> <p>That local ST612-MRSA-IV isolates described here share an uncommon <it>rpoB </it>genotype and a unique silent SNP suggests this clone may have undergone clonal expansion in hospitals in Cape Town. Further, the data suggest that these isolates may be related to rifampicin-resistant ST612-MRSA-IV previously described in South Africa and Australia.</p

Time for a culture change? Suboptimal compliance with blood culture standards at a district hospital in Cape Town

Background. The benchmark for contaminated blood cultures (BCs) is 3%. The South African (SA) guideline aims to optimise BC yield and reduce contamination. Data on BC collection practices in SA since the publication of the 2010 SA guideline are lacking.Objective. To evaluate compliance with the national guideline for the optimal use of BCs and determine the BC contamination rate at a local district hospital.Method. An audit of compliance with 22 BC standards was conducted at a district hospital in Cape Town, SA. Standards were evaluated by reviewing clinical and laboratory data and by a clinician questionnaire.Results. Of the 425 BCs reviewed, 12.5% had positive growth, and 4.5% grew contaminants. Only 33% of BC bottles contained the recommended fill volume of 8 - 10 mL, and 96.9% of patients had a single BC within a 24-hour period. Of all the BCs, only 7.8% had a combined blood volume of at least 20 mL. The yield of pathogens in BCs collected after antibiotic exposure was 4.9% compared with 7.5% for those cultures with no prior antibiotic exposure (p=0.3). The overall median needle-to-incubator transport time was 11 hours 25 minutes.Conclusion. The BC contamination rate was high and compliance with most standards was variable or not met. The findings may not be generalisable to other hospitals, and we recommend that each institution reviews its own BC practices. Recommendations made to hospital staff included a re-audit following implementation of these recommendations

Randomised controlled trial of early frenotomy in breastfed infants with mild-moderate tongue-tie

TRIAL DESIGN: A randomised, parallel group, pragmatic trial. SETTING: A large UK maternity hospital. PARTICIPANTS: Term infants <2 weeks old with a mild or moderate degree of tongue-tie, and their mothers who were having difficulties breastfeeding. OBJECTIVES: To determine if immediate frenotomy was better than standard breastfeeding support. INTERVENTIONS: Participants were randomised to an early frenotomy intervention group or a ‘standard care’ comparison group. OUTCOMES: Primary outcome was breastfeeding at 5 days, with secondary outcomes of breastfeeding self-efficacy and pain on feeding. Final assessment was at 8 weeks; 20 also had qualitative interviews. Researchers assessing outcomes, but not participants, were blinded to group assignment. RESULTS: 107 infants were randomised, 55 to the intervention group and 52 to the comparison group. Five-day outcome measures were available for 53 (96%) of the intervention group and 52 (100%) of the comparison group, and intention-to-treat analysis showed no difference in the primary outcome—Latch, Audible swallowing, nipple Type, Comfort, Hold score. Frenotomy did improve the tongue-tie and increased maternal breastfeeding self-efficacy. At 5 days, there was a 15.5% increase in bottle feeding in the comparison group compared with a 7.5% increase in the intervention group. After the 5-day clinic, 44 of the comparison group had requested a frenotomy; by 8 weeks only 6 (12%) were breastfeeding without a frenotomy. At 8 weeks, there were no differences between groups in the breastfeeding measures or in the infant weight. No adverse events were observed. CONCLUSIONS: Early frenotomy did not result in an objective improvement in breastfeeding but was associated with improved self-efficacy. The majority in the comparison arm opted for the intervention after 5 days

Impact of Xpert MTB/RIF for TB diagnosis in a primary care clinic with high TB and HIV prevalence in South Africa: a pragmatic randomised trial

Background: Xpert MTB/RIF is approved for use in tuberculosis (TB) and rifampicin-resistance diagnosis. However, data are limited on the impact of Xpert under routine conditions in settings with high TB burden. Methods and Findings: A pragmatic prospective cluster-randomised trial of Xpert for all individuals with presumptive (symptomatic) TB compared to the routine diagnostic algorithm of sputum microscopy and limited use of culture was conducted in a large TB/HIV primary care clinic. The primary outcome was the proportion of bacteriologically confirmed TB cases not initiating TB treatment by 3 mo after presentation. Secondary outcomes included time to TB treatment and mortality. Unblinded randomisation occurred on a weekly basis. Xpert and smear microscopy were performed on site. Analysis was both by intention to treat (ITT) and per protocol. Between 7 September 2010 and 28 October 2011, 1,985 participants were assigned to the Xpert (n = 982) and routine (n = 1,003) diagnostic algorithms (ITT analysis); 882 received Xpert and 1,063 routine (per protocol analysis). 13% (32/257) of individuals with bacteriologically confirmed TB (smear, culture, or Xpert) did not initiate treatment by 3 mo after presentation in the Xpert arm, compared to 25% (41/167) in the routine arm (ITT analysis, risk ratio 0.51, 95% CI 0.33–0.77, p = 0.0052). The yield of bacteriologically confirmed TB cases among patients with presumptive TB was 17% (167/1,003) with routine diagnosis and 26% (257/982) with Xpert diagnosis (ITT analysis, risk ratio 1.57, 95% CI 1.32–1.87, p<0.001). This difference in diagnosis rates resulted in a higher rate of treatment initiation in the Xpert arm: 23% (229/1,003) and 28% (277/982) in the routine and Xpert arms, respectively (ITT analysis, risk ratio 1.24, 95% CI 1.06–1.44, p = 0.013). Time to treatment initiation was improved overall (ITT analysis, hazard ratio 0.76, 95% CI 0.63–0.92, p = 0.005) and among HIV-infected participants (ITT analysis, hazard ratio 0.67, 95% CI 0.53–0.85, p = 0.001). There was no difference in 6-mo mortality with Xpert versus routine diagnosis. Study limitations included incorrect intervention allocation for a high proportion of participants and that the study was conducted in a single clinic. Conclusions: These data suggest that in this routine primary care setting, use of Xpert to diagnose TB increased the number of individuals with bacteriologically confirmed TB who were treated by 3 mo and reduced time to treatment initiation, particularly among HIV-infected participants

Ventilatory drive and the apnea-hypopnea index in six-to-twelve year old children

BACKGROUND: We tested the hypothesis that ventilatory drive in hypoxia and hypercapnia is inversely correlated with the number of hypopneas and obstructive apneas per hour of sleep (obstructive apnea hypopnea index, OAHI) in children. METHODS: Fifty children, 6 to 12 years of age were studied. Participants had an in-home unattended polysomnogram to compute the OAHI. We subsequently estimated ventilatory drive in normoxia, at two levels of isocapnic hypoxia, and at three levels of hyperoxic hypercapnia in each subject. Experiments were done during wakefulness, and the mouth occlusion pressure measured 0.1 seconds after inspiratory onset (P(0.1)) was measured in all conditions. The slope of the relation between P(0.1 )and the partial pressure of end-tidal O(2 )or CO(2 )(P(ET)O(2 )and P(ET)CO(2)) served as the index of hypoxic or hypercapnic ventilatory drive. RESULTS: Hypoxic ventilatory drive correlated inversely with OAHI (r = -0.31, P = 0.041), but the hypercapnic ventilatory drive did not (r = -0.19, P = 0.27). We also found that the resting P(ET)CO(2 )was significantly and positively correlated with the OAHI, suggesting that high OAHI values were associated with resting CO(2 )retention. CONCLUSIONS: In awake children the OAHI correlates inversely with the hypoxic ventilatory drive and positively with the resting P(ET)CO(2). Whether or not diminished hypoxic drive or resting CO(2 )retention while awake can explain the severity of sleep-disordered breathing in this population is uncertain, but a reduced hypoxic ventilatory drive and resting CO(2 )retention are associated with sleep-disordered breathing in 6–12 year old children

Trim28 Haploinsufficiency Triggers Bi-stable Epigenetic Obesity.

This is the final version of the article. It first appeared from Cell Press via http://dx.doi.org/10.1016/j.cell.2015.12.025More than one-half billion people are obese, and despite progress in genetic research, much of the heritability of obesity remains enigmatic. Here, we identify a Trim28-dependent network capable of triggering obesity in a non-Mendelian, "on/off" manner. Trim28(+/D9) mutant mice exhibit a bi-modal body-weight distribution, with isogenic animals randomly emerging as either normal or obese and few intermediates. We find that the obese-"on" state is characterized by reduced expression of an imprinted gene network including Nnat, Peg3, Cdkn1c, and Plagl1 and that independent targeting of these alleles recapitulates the stochastic bi-stable disease phenotype. Adipose tissue transcriptome analyses in children indicate that humans too cluster into distinct sub-populations, stratifying according to Trim28 expression, transcriptome organization, and obesity-associated imprinted gene dysregulation. These data provide evidence of discrete polyphenism in mouse and man and thus carry important implications for complex trait genetics, evolution, and medicine.This work was supported by funding from the Max-Planck Society, ERC (ERC-StG-281641), DFG (SFB992 “MedEp”; SFB 1052 “ObesityMechanisms”), EU_FP7 (NoE ”Epigenesys”; “Beta-JUDO” n° 279153), BMBF (DEEP), MRC (Metabolic Disease Unit - APC, SOR, GSHY, MRC_MC_UU_12012/1), Wellcome Trust (SOR, 095515/Z/11/Z) and the German Research Council (DFG) for the Clinical Research Center "Obesity Mechanisms" CRC1052/1 C05 and the Federal Ministry of Education and Research, Germany, FKZ, 01EO1001 (Integrated Research and Treatment Center (IFB) Adiposity Diseases

Bacterial Disease and Antimicrobial Susceptibility Patterns in HIV-Infected, Hospitalized Children: A Retrospective Cohort Study

The orginal version is available at www.plosone.orgBackground: Serious bacterial infections are a major source of morbidity and mortality in HIV-infected children. The spectrum of disease is wide, and responsible organisms vary according to setting. The use of antibiotic prophylaxis and the emergence of multi-drug resistant bacteria necessitate examination of responsible organisms and their antibiotic susceptibility. Methodology/Principal Findings: A retrospective cohort study of all HIV-positive pediatric admissions at an urban public sector hospital in Cape Town between January 2002 and June 2006 was conducted. Children between the ages of one month and nine years with laboratory confirmed HIV status, serious bacterial infection, and a hospital length of stay of 5 days or more, were eligible for inclusion. Organisms isolated from blood, urine, and cerebral spinal fluid cultures and their antimicrobial susceptibility were examined, and compared according to timing of isolation to distinguish nosocomial versus community-acquired. One hundred and forty-one children were identified (median age 1.2 years), 39% of whom were on antiretrovirals started before or during this hospitalization. Bacterial infections involved all organ systems, however pneumonia was most common (67%). S. pneumoniae and S. aureus were the most common gram positive and K. pneumoniae was the most common gram negative organism. K pneumoniae isolates were resistant to many first and second line antibiotics, and were all considered nosocomial. All S. aureus isolates were methicillin resistant, some of which were community-acquired. Conclusions/Significance: Bacterial infections are an important source of co-morbidity in HIV-infected children in resourcelimited settings. Clinicians should have a low threshold to initiate antibiotics in children requiring hospitalization. Broadspectrum antibiotics should be used judiciously. Clinicians caring for HIV-infected children should be cognizant of the most common organisms affecting such children, and of their local antimicrobial susceptibilities, when treating empirically for serious bacterial infections.Publisher's versio

Improved detection of Pneumocystis jirovecii in upper and lower respiratory tract specimens from children with suspected pneumocystis pneumonia using real-time PCR: a prospective study

<p>Abstract</p> <p>Background</p> <p><it>Pneumocystis </it>pneumonia (PCP) is a major cause of hospitalization and mortality in HIV-infected African children. Microbiologic diagnosis relies predominantly on silver or immunofluorescent staining of a lower respiratory tract (LRT) specimens which are difficult to obtain in children. Diagnosis on upper respiratory tract (URT) specimens using PCR has been reported useful in adults, but data in children are limited. The main objectives of the study was (1) to compare the diagnostic yield of PCR with immunofluorescence (IF) and (2) to investigate the usefulness of upper compared to lower respiratory tract samples for diagnosing PCP in children.</p> <p>Methods</p> <p>Children hospitalised at an academic hospital with suspected PCP were prospectively enrolled. An upper respiratory sample (nasopharyngeal aspirate, NPA) and a lower respiratory sample (induced sputum, IS or bronchoalveolar lavage, BAL) were submitted for real-time PCR and direct IF for the detection of <it>Pneumocystis </it><it>jirovecii</it>. A control group of children with viral lower respiratory tract infections were investigated with PCR for PCP.</p> <p>Results</p> <p>202 children (median age 3.3 [inter-quartile range, IQR 2.2 - 4.6] months) were enrolled. The overall detection rate by PCR was higher than by IF [180/349 (52%) vs. 26/349 (7%) respectively; p < 0.0001]. PCR detected more infections compared to IF in lower respiratory tract samples [93/166 (56%) vs. 22/166 (13%); p < 0.0001] and in NPAs [87/183 (48%) vs. 4/183 (2%); p < 0.0001]. Detection rates by PCR on upper (87/183; 48%) compared with lower respiratory tract samples (93/166; 56%) were similar (OR, 0.71; 95% CI, 0.46 - 1.11). Only 2/30 (6.6%) controls were PCR positive.</p> <p>Conclusion</p> <p>Real-time PCR is more sensitive than IF for the detection of <it>P. jirovecii </it>in children with PCP. NPA samples may be used for diagnostic purposes when PCR is utilised. Wider implementation of PCR on NPA samples is warranted for diagnosing PCP in children.</p