Application of normative documents for determination of biocidal activity of disinfectants and antiseptics dedicated to the medical area: a narrative review.

Disinfectants and antiseptics are important weapons to reduce the number of micro-organisms and thus limit the number of infections. Different methods of antimicrobial activity testing, often not standardized, without appropriate controls and not validated, are applied. To address these issues, several European Standards (EN) have been developed, describing the test methods to determine whether chemical disinfectants or antiseptic products have appropriate bactericidal, sporicidal, mycobactericidal or tuberculocidal activity; fungicidal or yeasticidal activity; or virucidal activity. In this narrative review, the 17 ENs concerning evaluation of the above-mentioned antimicrobial activity of preparations dedicated to the medical area are briefly reviewed, together with recent publications on this topic. Suspension and carrier tests have been performed in clean and dirty conditions simulating the medical area. In addition, a wide range of applications of these standards has been presented in the research of biocides for hand antisepsis, surfaces disinfection, including airborne disinfection as well as medical device and medical textile disinfection. The role of normative documents in the investigation of antimicrobial activity of disinfectants and antiseptics to limit infections has been underestimated. This narrative review aims to persuade researchers to conduct antimicrobial activity testing in line with validated ENs and highlights an existing gap in ongoing research. It also aims to raise awareness of the wide range of biocidal activity tests with standardized methods in the medical area. We also pay attention to the recently developed European Pharmacopoeia monography concerning the testing of bactericidal and fungicidal activity of antiseptics classified as medicinal products.

Application of metaphase HR-CGH and targeted Chromosomal Microarray Analyses to genomic characterization of 116 patients with mental retardation and dysmorphic features.

Recent advances in molecular cytogenetics enable identification of small chromosomal aberrations that are undetectable by routine chromosome banding in 5-20% of patients with mental retardation/developmental delay (MR/DD) and dysmorphism. The aim of this study was to compare the clinical usefulness of two molecular cytogenetic techniques, metaphase high-resolution comparative genomic hybridization (HR-CGH) and targeted array CGH, also known as Chromosomal Microarray Analysis (CMA). A total of 116 patients with unexplained mild to severe MR and other features suggestive of a chromosomal abnormality with apparently normal or balanced karyotypes were analyzed using HR-CGH (43 patients) and/or CMA (91 patients). Metaphase HR-CGH detected seven interstitial deletions (16.3%). Rare deletions of chromosomes 16 (16p11.2p12.1) and 8 (8q21.11q21.2) were identified. Targeted CMA revealed copy-number changes in 19 of 91 patients (20.8%), among which 11 (11.8%) were clinically relevant, 6 (6.5%) were interpreted as polymorphic variants and 2 (2.1%) were of uncertain significance. The changes varied in size from 0.5 to 12.9 Mb. In summary, our results show that metaphase HR-CGH and array CGH techniques have become important components in cytogenetic diagnostics, particularly for detecting cryptic constitutional chromosome imbalances in patients with MR, in whom the underlying genetic defect is unknown. Additionally, application of both methods together increased the detection rates of genomic imbalances in the tested groups.

Different-sized duplications of Xq28, including MECP2, in three males with mental retardation, absent or delayed speech, and recurrent infections.

In XY males, duplication of any part of the X chromosome except the pseudoautosomal region leads to functional disomy of the corresponding genes. We describe three unrelated male patients with mental retardation (MR), absent or delayed speech, and recurrent infections. Using high-resolution comparative genomic hybridization (HR-CGH), whole genome array comparative genomic hybridization (array CGH), fluorescent in situ hybridization (FISH), and multiplex ligation probe amplification (MLPA), we have identified and characterized two different unbalanced Xq27.3-qter translocations on the Y chromosome (approx. 9 and 12 Mb in size) and one submicroscopic interstitial duplication (approx. 0.3-1.3 Mb) involving the MECP2 gene. Despite the differences in size of the duplicated segments, the patients share a clinical phenotype that overlaps with the features described in patients with MECP2 duplication. Our data confirm previous observations that MECP2 is the most important dosage-sensitive gene responsible for neurologic development in patients with duplications on the distal part of chromosome Xq.

Neocentric small supernumerary marker chromosomes (sSMC)--three more cases and review of the literature.

Here we report on three new patients with neocentric small supernumerary marker chromosomes (sSMC) derived from chromosome 2, 13 and 15, respectively. The sSMC(13) and sSMC(15) had inverted duplicated shapes and the sSMC(2) a ring chromosome shape. All three cases were clinically severely abnormal. A review of the available sSMC literature revealed that up to the present 73 neocentric sSMC cases including these three new cases have been reported. Seven of these cases were not characterized morphologically; in the remainder, 80% had an inverted duplication, 17% a ring and 3% a minute shape. 81% of the reported neocentric sSMC carriers showed severe, 12% moderate and 8% no clinical abnormalities. In summary, we report three more neocentric sSMC cases, provide a review on all up to now published cases, highlight their special characteristics and compare them to centric sSMC.

Congenital anomalies and developmental delay in a boy with double chromosome 6 derived supernumerary marker.

The frequency of small supernumerary marker chromosomes has been estimated to approximately 0.45 per 1000 newborns. They are usually seen as single marker chromosomes in a mosaic state. Two cytogenetically identical markers have been observed only occasionally. We report on a boy, with congenital heart defect, neonatal hypotonia, hypogenitalism, delayed psychomotor development and mild dysmorphic facial features. The GTG karyotype performed on peripheral blood lymphocytes revealed a mosaic male karyotype with three cell lines. One cell line had a normal karyotype. In the other two either single or double chromosome 6 derived supernumerary markers were present, leading to partial trisomy or partial tetrasomy of chromosome 6, respectively.

Cytogenetic and molecular characterization of two isodicentric Y chromosomes.

We report the results of detailed molecular-cytogenetic studies of two isodicentric Y [idic(Y)] chromosomes identified in patients with complex mosaic karyotypes. We used fluorescence in situ hybridization (FISH) and polymerase chain reaction (PCR) to determine the structure and genetic content of the abnormal chromosomes. In the first patient, classical cytogenetics and FISH analysis with Y chromosome-specific probes showed in peripheral blood lymphocytes a karyotype with 4 cell lines: 45,X[128]/46,X,+idic(Y)(p11.32)[65]/47,XY,+idic(Y)(p11.32)[2]/47,X,+2idic(Y)(p11.32)[1]. No Y chromosome material was found in the removed gonads. For precise characterization of the Yp breakpoint, FISH and fiberFISH analysis, using a telomeric probe and a panel of cosmid probes from the pseudoautosomal region PAR1, was performed. The results showed that the breakpoint maps approximately 1,000 Kb from Ypter. The second idic(Y) chromosome was found in a boy with mild mental retardation, craniofacial anomalies, and the karyotype in lymphocytes 47,X,+idic(Y)(q11.23),+i(Y)(p10)[77]/46,X,+i(Y)(p10)[23]. To our knowledge, such an association has not been previously described. FISH and PCR analysis indicated the presence of at least two copies of the SRY gene in all analyzed cells. Using 17 PCR primers, the Yq breakpoint was shown to map between sY123 (DYS214) and sY121 (DYS212) loci in interval 5O in AZFb region. Possible mechanisms of formation of abnormal Y chromosomes and karyotype-phenotype correlations are discussed.

Frequency of Fra X syndrome among institutionalized mentally retarded males in Poland.

Results of cytogenetic studies, performed in a group of 201 institutionalized mentally retarded males, are presented. At least two cytogenetic methods for eliciting the Xq27.3 fragile site, recommended by the Fourth International Workshop on the Fra X Syndrome were used. A subgroup of 67 out of 201 studied males was also examined using molecular methods. In 6 (2.9%) males fra X syndrome was diagnosed. All cytogenetic positive results were confirmed by molecular analysis. Five patients had full expansion CGG repeats and one had both premutation and full mutation. Postulated frequency of fra X syndrome in Polish population being 0.2-0.4/1,000 males seems to be lower than it could be expected on the basis of previous literature data.

Clinical and molecular-cytogenetic studies in seven patients with ring chromosome 18.

We report the results of detailed clinical and molecular-cytogenetic studies in seven patients with ring chromosome 18. Classical cytogenetics and fluorescence in situ hybridization (FISH) analysis with the chromosome 18 painting probe identified five non-mosaic and two complex mosaic 46,XX,dup(18)(p11.2)/47,XX,dup(18)(p11.2),+r(18) and 46,XX,dup(18)(p11.32)/47,XX,dup(18)(p11.32),+r(18) cases. FISH analysis was performed for precise characterization of the chromosome 18 breakpoints using chromosome 18-specific short-arm paint, centromeric, subtelomeric, and a panel of fifteen Alu- and DOP-PCR YAC probes. The breakpoints were assessed with an average resolution of approximately 2.2 Mb. In all r(18) chromosomes, the 18q terminal deletions ranging from 18q21.2 to 18q22.3 ( approximately 35 and 9 Mb, respectively) were found, whereas only in four cases could the loss of 18p material be demonstrated. In two cases the dup(18) chromosomes were identified as inv dup(18)(qter-->p11.32::q21.3-->qter) and inv dup(18)(qter-->p11.32::p11.32-->p11.1: :q21.3-->qter)pat, with no evidence of an 18p deletion. A novel inter-intrachromatid mechanism of formation of duplications and ring chromosomes is proposed. Although the effect of "ring instability syndrome" cannot be excluded, the phenotypes of our patients with characteristic features of 18q- and 18p- syndromes are compared and correlated with the analyzed genotypes. It has been observed that a short neck with absence of cardiac anomalies may be related to the deletion of the 18p material from the r(18) chromosome.

Analysis of unstable DNA sequence in FMR1 gene in Polish families with fragile X syndrome.

The unstable DNA sequence in the FMR1 gene was analyzed in 85 individuals from Polish families with fragile X syndrome in order to characterize mutations responsible for the disease in Poland. In all affected individuals classified on the basis of clinical features and expression of the fragile site at X(q27.3) a large expansion of the unstable sequence (full mutation) was detected. About 5% (2 of 43) of individuals with full mutation did not express the fragile site. Among normal alleles, ranging in size from 20 to 41 CGG repeats, allele with 29 repeats was the most frequent (37%). Transmission of premutated and fully mutated alleles to the offspring was always associated with size increase. No change in repeat number was found when normal alleles were transmitted.

[Prenatal diagnosis of chromosomal aberrations in first trimester pregnancy--methods for cytogenetic analysis of the trophoblast]. / Diagnostyka prenatalna aberracji chromosomowych w i trymestrze ciazy--metody analizy cytogenetycznej trofoblastu.

Prenatal diagnosis of chromosomal abnormalities by chorionic villus (CVS) in the first trimester of pregnancy is presented. Trophoblast biopsy techniques, methods of cytogenetic analysis of this tissue as well as technical and diagnostic problems in fetal karyotyping are described.

[Results of 1043 prenatal cytogenetic studies: retrospective study in the context of applicability of interphase FISH in prenatal diagnosis]]. / Retrospektywna analiza 1043 wyników prenatalnych badan cytogenetycznych w kontekscie przydatnosci diagnostycznej FISH interfazowej.

OBJECTIVES: The risk of aneuploidy in a fetus is the main reason for referral in approximately 80% of prenatal studies. Recently, a new method for rapid detection of the most frequent aneuploidies affecting chromosomes 13, 18, 21, X and Y has been developed. Fluorescence in situ hybridisation (FISH) on uncultured fetal cells with probes specific for these chromosomes has been described which enables diagnosing aneuploidies within 24 to 48 hours. The purpose of the study was evaluation of clinical utility of this new method in prenatal diagnostics. MATERIALS AND METHODS: Retrospective analysis of the results of 1043 prenatal cytogenetic studies performed with conventional banding methods was done. Number and type of chromosomal abnormalities found in different categories of indications with special emphasis on aberrations undetectable by FISH were analysed. RESULTS: Chromosomal aberrations were found in 4.7% studies. The frequency of aneuploidies was 1.8% accounting for 35.8% of all diagnosed chromosomal abnormalities. In the group of 854 studies performed for elevated risk of aneuploidy it accounted for 60.7% (17/28) abnormalities. All other aberrations could not be detected by FISH with probes for most frequent aneuploidies. Among them, there were 6 unbalanced: del (8) and pseudic (15) with known abnormal phenotype and 4 marker chromosomes with unknown clinical consequences. Three other abnormalities were balanced but familial origin of them was documented. CONCLUSIONS: A karyotype using classical banding methods should be performed whatever the indication of prenatal study is. It is the only fully informative method able to detect all chromosomal abnormalities. Interphase FISH assay must be considered as a complementary procedure to fetal karyotype analysis as it is designed only for aneuploidy identification. However it may be a very useful method for rapid diagnosis in specific clinical conditions especially in the cases of high risk of aneuploidy.

[Prenatal diagnosis of chromosomal aberrations in the second trimester of pregnancy--analysis of problems and diagnostic difficulties]. / Diagnostyka prenatalna aberracji chromosomalnych w II trymestrze ciazy--analiza problemów i trudnosci diagnostycznych.

Laboratory problems and diagnostic difficulties are analysed in 1135 prenatal cytogenetic examinations of amniotic fluid cells in the Genetics Laboratory of the Mother and Child Institute. The effectiveness and the reliability of the method for the purposes of prenatal diagnosis of chromosomal aberrations were evaluated. They were comparable to those reported from other similar centres in the world.

[Cytogenetic studies in clinical diagnosis --analysis of 1611 examination results]. / Badania cytogenetyczne w diagnostyce klinicznej--analiza 1611 wyników badan.

An analysis of the frequency and type of chromosome aberrations found in 1611 cytogenetic studies has been done in relation to the clinical indications. The most frequent reason for referral was: MCM/MR syndrome (15.4% cases), suspicion of sex chromosome abnormality (13.1%), Down syndrome (11.2%) and reproductive wastage (10.6%). The incidence of abnormal karyotypes in these groups of patients was 10.1%, 24.2%, 87.9% and 9.3% per couple, respectively. The lowest percentage of chromosome aberrations was found among patients with nonspecific mental retardation (2.2%) and in the families who lost a baby with MCM Syndrome (3.3% per couple). On the whole, abnormal karyotype was found in 364 patients (22.6%). Among 62 families identified by the affected child with, structural chromosome aberration in 24 families (38.7%) the abnormality was familial in the origin.

[Marker chromosomes as a product of familial translocation (11;22) identified with molecular cytogenetic methods]. / Marker chromosomowy jako produkt rodzinnej translockacji (11;22) rozpoznanej metodami cytogenetyki molekularnej.

A reciprocal constitutive 11;22 translocation is the most frequent, non Robertsonian translocation in man. We describe a case of partial trisomy 11q and 22q in a child with facial dysmorphy, hypotonia, heart failure, cryptorchism and psychomotor retardation. A marker chromosome was found in this child. Chromosome analysis with the fluorescence in situ hybridization, FISH technique showed that this marker chromosome was the product of 3:1 mejotic segregation of maternal (11;22) balanced translocation. Routine cytogenetic problems with identification of marker chromosomes can now successfully be solved with the FISH technique. The presented case clearly demonstrates the diagnostic usefulness of this newest method of cytogenetic analysis.

[Reliability of cytogenetic and clinical diagnostic studies. Results of analyzing 1621 pre- and postnatal studies]. / Wiarygodnosc badan cytogenetycznych w diagnostyce klinicznej. Wyniki analizy 1621 badan pre- i postnatalnych.

Systematic quality assessment of cytogenetic studies is very important for maintaining high standards of diagnostic testing and adequate genetic service. Results of quality assessment for 714 pre- and 907 postnatal cytogenetic studies performed in the Genetic Department of the National Research Institute of Mother and Child are presented. The assessment included: reporting time, quality of chromosome preparations, reliability and success rate of the studies. Mean reporting time for blood samples was 20 days and for amniotic fluid 23 days. Chromosome banding quality was adequate to reasons for referral in 86.9% of blood samples and in 100% of amniotic fluids. The success rate for pre- and postnatal studies was 99.4 and 96.6%, respectively. The need to develop a quality assessment scheme for clinical cytogenetics in Poland is discussed.

[Use of molecular cytogenetic techniques for establishing the origin of chromosome markers in patients with Turner phenotype]. / Zastosowanie technik cytogenetyki molekularnej w ustalaniu pochodzenia chromosomów markerowych u chorych z fenotypem Turnera.

Results of marker chromosome identification using the FISH technique are presented. The origin of markers was determined in 11 patients with mosaic karyotype 45,X/46,X,mar. Using probes specific for X and Y chromosomes, we demonstrated that in 8 patients the marker originated from the X chromosome and in 3 cases it was an abnormal Y chromosome. These 3 patients were identified as at high risk for developing gonadoblastoma and prophylactic gonadectomy was performed. Results of these studies show that the FISH technique is a very useful method in the diagnosis of sex chromosome abnormalities.

[Partial trisomy of chromosome 13--diagnosis confirmed with the FISH in situ hybridization technique]. / Czesciowa trisomia chromosomu 13--rozpoznanie potwierdzone technika hybrydyzacji in situ FISH.

The case of a 1.5 year old girl with clinical traits of craniofacial dysmorphy, hypotonia, polydactyly and moderate mental retardation is presented. Routine cytogenetic study revealed the presence of a large additional chromosomal fragment associated with the nucleolus organizing region on one of chromosomes 13. The banding pattern suggested the additional fragment was a part of the long arm of this chromosome. The set of clinical symptoms was only partly consistent with those characteristic for trisomy 13q2 and 3. Application of the FISH technique with a chromosome 13 specific library enabled final confirmation of the origin of the extra chromosome fragment from the long arm of chromosome 13. The presented case proves the usefulness of the FISH technique for the diagnosis of chromosomal aberrations and for adequate clinical interpretation of cytogenetic results.