- Volume 67, Issue 7, 2018
Volume 67, Issue 7, 2018
- Antimicrobial Resistance
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Antifungal susceptibility and virulence of Candida parapsilosis species complex: an overview of their pathogenic potential
Raimunda Sâmia Nogueira Brilhante, Jamille Alencar Sales, Maria Lucilene Queiroz da Silva, Jonathas Sales de Oliveira, Lucas de Alencar Pereira, Waldemiro Aquino Pereira-Neto, Rossana de Aguiar Cordeiro, José Júlio Costa Sidrim, Débora de Souza Collares Maia Castelo-Branco and Marcos Fábio Gadelha RochaPurpose. Antifungal resistance and several putative virulence factors have been associated with the pathogenicity of the Candida parapsilosis species complex. The objective of this study was to evaluate the antifungal susceptibility, the production of virulence factors and the pathogenicity of the C. parapsilosis complex.
Methodology. Overall, 49 isolates of C. parapsilosis sensu stricto, 19 C. orthopsilosis and nine C. metapsilosis were used. The planktonic and biofilm susceptibility to fluconazole, itraconazole, voriconazole, amphotericin B and caspofungin was assessed using a broth microdilution assay. Finally, the production of biofilm and hydrolytic enzymes and the fungal pathogenicity against Caenorhabditis elegans were investigated.
Results/Key findings. Overall, one C. orthopsilosis was resistant to caspofungin and susceptible-dose-dependent to itraconazole, the other two C. orthopsilosis were susceptible-dose-dependent to fluconazole and itraconazole, and one C. metapsilosis was susceptible-dose-dependent to azoles. A total of 67.5 % of the isolates were biofilm producers. Amphotericin B and caspofungin caused the greatest reduction in the metabolic activity and biomass of mature biofilms. Phospholipase and protease production was observed in 55.1 % of C. parapsilosis sensu stricto, 42.1 % of C. orthopsilosis and 33.3 % of C. metapsilosis isolates. Moreover, 57.9 % of C. orthopsilosis and 20.4 % of C. parapsilosis sensu stricto isolates were β-haemolytic, and all C. metapsilosis were α-haemolytic. Finally, the C. parapsilosis complex caused high mortality of C. elegans after 96 h of exposure.
Conclusion. These results reinforce the heterogeneity of these cryptic species for their antifungal susceptibility, virulence and pathogenic potential, emphasizing the relevance of monitoring these emerging pathogens.
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High diversity in SCCmec elements among multidrug-resistant Staphylococcus haemolyticus strains originating from paediatric patients; characterization of a new composite island
Purpose. Staphylococcus haemolyticus has emerged as a highly antimicrobial-resistant healthcare-associated pathogen, in particular for patients admitted to neonatal intensive care. The objective of this study was to study the nature of SCCmec types among MDR-SH strains isolated from paediatric patients.
Methodology. S. haemolyticus strains (n=60) were isolated from paediatric patients. Antibiotic resistance patterns were established using the disk agar diffusion and micro-broth dilution methods. SCCmec typing was performed using whole-genome sequencing (WGS) and an additional PCR analysis.
Results. All S. haemolyticus isolates demonstrated multidrug resistance. Using WGS, various novel mec types and combinations of SCCmec types were found, including a new composite island [SCCmec type V (Vd)+SCC cad/ars/cop] comprising 30 % of the strains. SCCmec type V was identified in 23 % of the isolates. A combination of the mecA gene enclosed by two copies of IS431 and absence of the mecRI and ccr genes was identified in 11 strains. In total, mecA regulatory genes were absent in all SH isolates used in this study.
Conclusion. A high diversity of SCCmec elements with the prevalence of a new composite island was determined among MRSH strains. The structure of the composite island represented by MDR-SH strains in this study, in combination with the presence of a restriction-modification system type III, is described for the first time in this study. The presence of an 8 bp direct repeat (DR) and the sequences flanking the DR may support the integration of the mecA gene complex as a composite transposon (IS431-mecA-IS431) independently from recombinase genes.
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Relevance of antifungal penetration in biofilm-associated resistance of Candida albicans and non-albicans Candida species
More LessThe role of penetration limitation in Candida biofilm-associated antifungal resistance remains unclear. Most of the previous work has been done on Candida albicans, although non-albicans (NAC) species are also implicated in invasive candidiasis and the biofilm matrix has been shown to vary amongst different species. Only a few studies have evaluated clinical isolates. This study aimed to determine the relevance of penetration limitation in the antifungal resistance of biofilms formed by C. albicans and NAC clinical isolates, using an agar disk diffusion assay. The penetration of posaconazole and amphotericin B through the biofilms was significantly reduced. Fluconazole, voriconazole and caspofungin showed a superior penetration capacity in C. albicans, Candida tropicalis and Candida parapsilosis biofilms, but exhibited inter-species and strain/isolate variation. Candida krusei biofilms were the most resilient to antifungal permeation. All of the antifungal drugs failed to kill the biofilm cells, independent of penetration, suggesting that the other factors contribute markedly to the recalcitrance of the biofilms.
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KPC-2 producing ST101 Klebsiella pneumoniae from bloodstream infection in India
More LessThis study characterizes KPC-2 producing Klebsiella pneumoniae belonging to ST101. Whole genome sequencing using the Ion Torrent PGM platform with 400 bp chemistry was performed. bla KPC-2 was found on an IncFIIK plasmid associated with ISKpn6 and ISKpn7 without Tn4401. This is the first report of KPC-2 K. pneumoniae from bacteremia in India. The isolate also coded for other resistance genes such as aadA1, aadA2, armA, aac(3)-Ild, aac(6′)-Ild for aminoglycoside; bla SHV-11 , bla TEM-1B , bla OXA-9, for β-lactams and aac(6′)-Ild, oqxA, oqxB, qnrB1 for fluoroquinolones. It belonged to the K17 capsular type. India is endemic to New Delhi metallo-β-lactamase and OXA48-like carbapenemases and K. pneumoniae carbapenemase (KPC) is seldom reported. With high rates of carbapenem resistance, emergence of KPC in India will challenge patient management. The isolate was susceptible to colistin. The patient had a fatal outcome.
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Occurrence and characterization of extended-spectrum cephalosporin-resistant Enterobacteriaceae in healthy household dogs in Greece
Extended-spectrum cephalosporin- and/or carbapenem-resistant (ESCR and/or CarbR) Enterobacteriaceae constitute a public health hazard because of limited treatment options and are endemic among humans in Greece. Recently, ESCR and CarbR Enterobacteriaceae have been increasingly isolated from companion animals, stressing their potential role as a reservoir for humans. However, the presence of ESCR bacteria in companion animals within Greek households has not been determined yet. Genes conferring the ESCR and CarbR phenotype were detected among canine isolates and their chromosomal or plasmid location was determined. Standard methods were applied for plasmid characterization. The clonal relatedness of the recovered isolates was examined by multilocus sequence typing (MLST). Here, we report the first findings on the presence of ESCR Enterobacteriaceae in healthy Greek dogs. ESCR Escherichia coli isolates were associated with different sequence types (STs), including the human pandemic ST131 clone. The occurrence of human-related ESBL/pAmpC genes, plasmid types and/or strain STS in this animal reservoir suggests possible bilateral transmission.
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- Clinical Microbiology
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Can dalbavancin be used as a catheter lock solution?
Purpose. The new lipoglycopeptide dalbavancin has only been approved for acute bacterial skin and skin structure infections. However, its alternative use as a catheter lock solution could facilitate the conservative management of catheter-related bloodstream infection. Our objective was to assess the stability and activity of dalbavancin alone and in combination with heparin against methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-resistant Staphylococcus epidermidis (MRSE) biofilms. We also compared the results with those obtained with vancomycin alone and in combination with heparin.
Methodology. We used a 96-well plate in vitro model based on 24 h biofilms of MRSA and MRSE (ATCC 43300, ATCC 35984 and one clinical strain of each). The biofilms were exposed to dalbavancin (0.128 mg ml−1) and vancomycin (5 mg ml−1) alone and in combination with heparin (60 IU). The median percentage reductions in metabolic activity, biomass, bacterial load, and cell viability for each solution were compared.
Results. Dalbavancin combined with heparin significantly reduced the median [interquartile range (IQR)] percentage of metabolic activity in MRSA biofilms compared with vancomycin [90.0 % (70.4–92.9 %) versus 35.0 % (14.8–59.6 %), P=0.006]. For the remaining variables studied, the combination was not inferior to vancomycin for MRSA and MRSE.
Conclusions. Dalbavancin proved to be active against MRSA and MRSE biofilms. The combination of dalbavancin with heparin is a promising catheter lock solution that has the advantage of locking the catheter at home for 7 days.
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Regulation of gene expression of hcp, a core gene of the type VI secretion system in Acinetobacter baumannii causing respiratory tract infection
More LessPurpose. The objective of the current study was to investigate whether hcp plays a role in the process of Acinetobacter baumannii infection and to examine clinically relevant factors that may affect hcp expression.
Methodology. Seventy-seven A. baumannii isolates from patients with a respiratory infection at the Second Affiliated Hospital and Yuying Childrens Hospital of Wenzhou Medical University (Wenzhou, China) were included in this study. PCR was performed to screen for the presence of hcp. Quantitative real time polymerase chain reaction (qRT-PCR) was carried out to examine the expression of hcp.
Results. A total of 77.9 % (60 of 77) of the A. baumannii clinical isolates possessed the hcp gene. Expression of hcp was found to be strain-specific and associated with the infection status. Higher gene expression of hcp was found for invasive A. baumannii isolates causing an infection relative to the colonization group, and for the same strain at a post-infection status compared with that prior to infection. Acid environment was also found to be a trigger of hcp gene expression.
Conclusion. The type VI secretion system and hcp predominate in A. baumannii causing respiratory infections. Expression of hcp is regulated by the infection status and acid environment, and might play a role in the process of triggering infection by the colonizer.
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Effect of respiratory Achromobacter spp. infection on pulmonary function in patients with cystic fibrosis
More LessPurpose. Cystic fibrosis (CF) patients are susceptible to infection with Achromobacter spp., although its clinical significance remains controversial. The aim of this study was to investigate the clinical impact of infection with Achromobacter spp. in CF patients.
Methods. CF outpatients with multiple sputum cultures and follow-up lung function tests were assigned to the case group (infected with Achromobacter spp.) or the control group (never infected with Achromobacter spp.) according to the isolation of Achromobacter spp. The Achromobacter spp. group included two subgroups, taking into consideration whether the isolation of Achromobacter spp. was intermittent or chronic. Baseline lung function tests and longitudinal behaviour were examined in relation to Achromobacter spp. status.
Results. A total of 190 CF patients were treated from January 2003 to December 2015 in the CF unit and 21 (11 %) had at least one positive culture for Achromobacter spp. Of these, 11/21 (52.4 %) patients were chronically infected with Achromobacter spp. An analysis of changes during follow-up showed the annual rate of FEV1 decline: −2.3±1.6 % in the Achromobacter spp. group compared to −1.1±0.9 % (P=0.02) in the control group. The chronically infected group also had a significantly greater decline in FEV1 compared to the control group (−2.9±1.9 vs −1.1±0.9; P=0.04). The mean number of annual pulmonary exacerbations during the study period was significantly higher in the case group (1.9±0.9 vs 1.1±0.8; P=0.03).
Conclusions. The Achromobacter spp. status in CF shows a trend towards more severe airflow obstruction and an association with accelerated decline in lung function parameters.
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Emergence and nosocomial spread of ST11 carbapenem-resistant Klebsiella pneumoniae co-producing OXA-48 and KPC-2 in a regional hospital in Taiwan
Purpose. Carbapenem-resistant Klebsiella pneumoniae (CRKP) has emerged as a major challenge for global healthcare systems. The objectives of this study were to determine the nosocomial spread of CRKP clones and analyse the molecular characteristics of CRKP in our hospital.
Methodology. Ninety-eight non-duplicated clinical CRKP isolates were collected from March 2014–June 2015. Clinical, demographic and microbiological data of patients with CRKP were reviewed. Pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing were applied to investigate the genetic relationship between the 98 isolates. Antibiotic resistance genes were identified by conventional PCR-sequencing.
Results. PFGE patterns were grouped into 26 clusters. Two main PFGE clusters were identified: L (53 isolates, belonging to ST11) and N (11 isolates, belonging to ST11). The most dominant ST was ST11 (79 %, 77/98), followed by ST273 (5 %, 5/98). KPC-2 (n=82) was the predominant carbapenemase followed by OXA-48 (n=64). Fifty isolates (51 %, 50/98) harboured bla KPC-2 and bla OXA-48 simultaneously, and three of these isolates were detected with the third carbapenemase genes (bla IMP-8 or bla VIM-1).
Conclusion. The clonal spread of K. pneumoniae ST11 expressing OXA-48, KPC-2 and CTX-M-14 β-lactamases was the cause of an outbreak of CRKP. To the best of our knowledge, a single strain harbouring A-, B- and D-class carbapenemase genes has not previously been identified. There is a high prevalence of plasmid-encoded KPC-2- and OXA-48-producing CRKP in our hospital; most isolates were members of ST11, which may be representative of a high-risk CRKP clone disseminating in central Taiwan.
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Nalidixic acid surrogate test for susceptibility to ciprofloxacin in Salmonella. Revisiting the question
We investigated the reliability of nalidixic acid (NA) susceptibility as a marker of ciprofloxacin susceptibility in Salmonella, analysing 302 stool isolates. NC53 of the MicroScan system was used for NA susceptibility tests and the E-test was used for ciprofloxacin susceptibility tests. Among the isolates, 178 (58.9 %) were serogroup B, 74 (24.5 %) were serogroup D, 27 (8.9 %) were serogroup C and 23 (7.6 %) were from other minor serogroups. Globally, susceptibility to NA correctly predicted the susceptibility of Salmonella to ciprofloxacin, with a sensitivity of 81.5 %, a specificity of 97.6 %, and positive and negative predictive values of 88 and 96 %, respectively. However, there were differences among the serogroups in terms of sensitivity (P<0.001) and positive predictive values (P=0.013). NA is a reliable marker for serogroup D, but not for serogroups B or C. According to these findings, NA susceptibility measured with the MicroScan system can be used as a marker of ciprofloxacin resistance in some serogroups in our setting.
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Methicillin-resistant Staphylococcus argenteus misidentified as methicillin-resistant Staphylococcus aureus emerging in western Sweden
Two strains included in a whole-genome sequencing project for methicillin-resistant Staphylococcus aureus (MRSA) were identified as non-Staphylococcus aureus when the sequences were analysed using the bioinformatics software ALEX (www.1928diagnostics.com, Gothenburg, Sweden). Sequencing of the sodA gene of these strains identified them as Staphylococcus argenteus. The collection of MRSA in western Sweden was checked for additional strains of this species. A total of 18 strains of S. argenteus isolated between 2011 and December 2017 were identified.
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Tigecycline resistance among Klebsiella pneumoniae isolated from febrile neutropenic patients
More LessFebrile neutropenic patients are at a high risk of life-threatening bacterial infections. Tigecycline was developed to treat multidrug-resistant isolates, however resistance to tigecycline in Klebsiella pneumoniae has been reported. Here, we investigated tigecycline resistance among K. pneumoniae isolated from febrile neutropenic patients admitted to Hematology ICU, Egypt. Out of 75 enrolled febrile neutropenic patients, 48 cases showed bacteriologically confirmed infection. The majority of cases were infected with K. pneumoniae, of which nine were tigecycline non-susceptible. Expression levels of the efflux pump genes acrB and oqxB and their regulatory genes ramA and rarA were analysed. Six isolates had overexpression of the four efflux-related genes while one showed baseline expression. This study emphasizes the importance of growing tigecycline resistance in K. pneumoniae infecting febrile neutropenic patients. Concerning the mechanism of resistance, it was clear that the ramA gene plays the major role, although alternative resistance mechanisms may also exist.
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- Disease, Diagnosis and Diagnostics
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Detection of toxigenic Clostridium difficile colonization in patients admitted to the hospital for chemotherapy or haematopoietic cell transplantation
Increasing evidence suggests that asymptomatic carriers are an important source of healthcare-associated Clostridium difficile infection. However, it is not known which test for the detection of C. difficile colonization is most sensitive in patients with haematological malignancies. We performed a prospective cohort study of 101 patients with haematological malignancies who had been admitted to the hospital for scheduled chemotherapy or haematopoietic cell transplantation. Each patient provided a formed stool sample. We compared the performance of five different commercially available assays, using toxigenic culture as the reference method. The prevalence of toxigenic C. difficile colonization as determined by toxigenic culture was 14/101 (14 %). The Cepheid Xpert PCR C. difficile/Epi was the most sensitive test for the detection of toxigenic C. difficile colonization, with 93 % sensitivity and 99 % negative predictive value. Our findings suggest that the Xpert PCR C. difficile/Epi could be used to rule out toxigenic C. difficile colonization in this population.
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- Microbial Ecology and Health
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Lactobacillus plantarum ameliorates tumour necrosis factor-induced bacterial translocation in Caco-2 cells by regulation of TLR4 expression
More LessPurpose. Translocation of bacteria across the intestinal barrier is important in the pathogenesis of systemic sepsis. In inflammatory conditions, commensal bacteria exploit transcytotic pathways to cross the intestinal epithelium in a TLR4-dependent manner. The aim of this study was to test the hypothesis that Lactobacillus plantarum ameliorates tumour necrosis factor-induced bacterial translocation by regulation of Toll-like receptor-4 expression.
Methodology. L. plantarum strains were investigated to determine their capacity to inhibit the initial adhesion of Escherichia coli B5 to Caco-2 cells. The inhibitory effects of L. plantarum on TNF-α-induced E. coli B5 translocation across Caco-2 cells were studied. Barrier function and integrity were simultaneously assessed by transepithelial electrical resistance, HRP permeability, LDH release and distribution of tight junctional proteins. Expression of TLR4 was assessed by RT-PCR.
Results/Key findings. Pretreatment of monolayers with L. plantarum L2 led to a significant decrease in E. coli B5 adhesion and cell internalization (P<0.01). Exposure to TNF-α for six hours caused a significant increase in E. coli B5 translocation across Caco-2 cells, which was uncoupled from increases in paracellular permeability and disruption of tight junction proteins. Manipulations that induced bacterial translocation were associated with a marked increase in TLR4 mRNA expression and IL-8 secretion. L. plantarum L2 significantly abrogated TNF-α-induced bacterial translocation of E. coli B5, and also downregulated expression of TLR4 and IL-8 in intestinal epithelial cells.
Conclusion. Live L. plantarum L2 can inhibit TNF-α-induced transcellular bacterial translocation via regulation of TLR4 expression.
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- Microbial Epidemiology
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Stenotrophomonas maltophilia isolated from patients exposed to invasive devices in a university hospital in Argentina: molecular typing, susceptibility and detection of potential virulence factors
Purpose. The aim of this work was to investigate the presence of selected potential virulence factors, susceptibility and clonal relatedness among 63 Stenotrophomonas maltophilia isolates recovered from patients exposed to invasive devices in a university hospital in Argentina between January 2004 and August 2012.
Methodology. Genetic relatedness was assessed by enterobacterial repetitive intergenic consensus PCR (ERIC-PCR) and pulsed-field gel electrophoresis (PFGE). Isolates were characterized by antimicrobial resistance, the presence and/or expression of potential virulence determinants, and virulence in the Galleria mellonella model.
Results/Key findings. ERIC-PCR generated 52 fingerprints, and PFGE added another pattern. Resistance to trimethoprim–sulfamethoxazole (6.35 %), levofloxacin (9.52 %) and ciprofloxacin (23.80 %) was detected. All isolates were susceptible to minocycline. All isolates were lipase, protease and siderophore producers, while all but Sm61 formed biofilms. However, 11/63 isolates did not amplify the major extracellular protease-coding gene (stmPr1). Sm61 is an stmPr1-negative isolate, and showed (as did Sm13 and the reference strain K279a) strong proteolysis and siderophore production, and high resistance to hydrogen peroxide. The three isolates were virulent in the G. mellonella model, while Sm10, a low-resistance hydrogen peroxide stmPr1-negative isolate, and weak proteolysis and siderophore producer, was not virulent.
Conclusion. This is the first epidemiological study of the clonal relatedness of S. maltophilia clinical isolates in Argentina. Great genomic diversity was observed, and only two small clusters of related S. maltophilia types were found. Minocycline and trimethoprim–sulfamethoxazole were the most active agents. S. maltophilia virulence in the G. mellonella model is multifactorial, and further studies are needed to elucidate the role of each potential virulence factor.
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Molecular characterization of serogroup 19 Streptococcus pneumoniae in the Czech Republic in the post-vaccine era
Purpose. The aim of this study was to characterize serogroup 19 isolates resistant to macrolides and/or penicillin found among pneumococci recovered from cases of invasive and respiratory tract disease in the Czech Republic in 2014.
Methods. Pneumococcal isolates of serotypes 19A (n=26) and 19F (n=10) that were non-susceptible to penicillin and/or macrolides and had been collected in 2014 were analysed using multi-locus sequence typing (MLST). Four isolates representing the major clones were subjected to whole-genome sequencing (WGS).
Results. The penicillin-susceptible macrolide-resistant isolates of serotype 19A were mainly associated with sequence type (ST) 416 belonging to clonal complex (CC) 199, and the penicillin-resistant isolates were of serotype 19F belonging to ST1464 (CC 320). WGS revealed the presence of pilus 1, in association with pilus 2, in serotype19F isolates belonging to CC 320. Another adhesin, pneumococcal serine-rich protein (PsrP), was only present in serotype 19A isolates of ST416. Analysis of the penicillin-binding proteins (PBPs) of serotype 19F penicillin-resistant isolates (ST1464 and ST271) performed on PBP1a, 2b and 2x identified a large number of mutations in comparison to the reference strain, R6. Both isolates contained a unique PBP profile; however, they were highly similar to PBP sequences of the Taiwan19F-14 reference strain. The Pbp2b sequences of both 19F isolates showed the lowest similarity to those of the Taiwan19F-14 strain (91 % similarity), while they were also found to be distantly related to each other (94 % similarity).
Conclusions. WGS revealed specific virulence factors in antibiotic-resistant pneumococcal clones that spread rapidly in the post-vaccine era in the Czech Republic.
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- One Health
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An accidental laboratory exposure to Brucella melitensis: the prospective post-exposure management and a detailed investigation into the nature of the exposure
More LessOur aim was to prospectively manage 22 Brucella-exposed individuals and identify the lapses in laboratory practices that lead to the exposure. The exposed individuals were risk-stratified, assessed for post-exposure prophylaxis (PEP), counselled to self-monitor symptoms and followed-up with three serology tests. Staff laboratory practices were recorded. Ten out of 13 high-risk individuals received PEP within 48 h of exposure. Compliance with PEP and serology monitoring was 90 and 96 %, respectively. No brucellosis cases were documented. A single handler manipulated the Brucella isolate on the open bench. Movement of the isolate was tracked in detail, highlighting various points of laboratory non-conformance. Early PEP intervention is effective in preventing acquired brucellosis. Our pragmatic post-exposure management achieved high PEP and serology compliance. We experience first-hand how regular staff engagement motivated PEP adherence and interval blood sampling attendance. The enforcement of practical strategies and safety practices was also implemented without compromising our laboratory processing times.
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Volumes and issues
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Volume 73 (2024)
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Volume 72 (2023 - 2024)
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Volume 71 (2022)
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Volume 70 (2021)
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Volume 69 (2020)
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Volume 68 (2019)
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Volume 67 (2018)
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Volume 66 (2017)
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Volume 65 (2016)
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Volume 64 (2015)
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Volume 63 (2014)
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Volume 3 (1970)
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Volume 2 (1969)
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Volume 1 (1968)