59 research outputs found
Biolog identification of non-sorbitol fermenting bacteria isolated on E. coli O157 selective CT-SMAC agar
E. coli O157:H7 is recognised as an important human pathogen world-wide and has been associated with diseases such as
haemorrhagic colitis (HC), haemolytic uraemic syndrome (HUS) and thrombotic thrombocytopaenic purpura (TTP). Accurate
laboratory detection of E. coli O157:H7 is important for diagnostic purposes and to justify epidemiological data on E.
coli O157:H7. A well-known phenotypic characteristic of E. coli O157:H7 bacteria is their inability to ferment sorbitol. This
characteristic is often used to isolate these organisms from food and water using selective agar medium such as SMAC. However,
the high number of false positive results obtained by a number of researchers when selectively screening for E. coli O157:
H7 on CT-SMAC has prompted an investigation to determine which other sorbitol-negative bacteria also grow on CT-SMAC.
The agar medium used for the investigation consisted of Sorbitol MacConkey agar (SMAC) supplemented with Cefiximetellurite
(CT). All sorbitol-negative colonies obtained from CT-SMAC, after selective enrichment and IMS were identified
using the Biolog microbial identification system. The majority of sorbitol-negative isolates identified were Burkholderia,
Pseudomonas, Vibrio and Aeromonas spp. Only two E. coli O157:H7 isolates were identified with Biolog and confirmed with
a polymerase chain reaction (PCR) specific for the shiga toxin 1 (Stx1) genes and with O157 and H7 antisera. The inability
of the CT-SMAC agar medium to specifically select for E.coli O157:H7 was confirmed by the results of this study. These
observations call for further improvement of affordable methods for the selective isolation of E. coli O157:H7 in the presence
of large numbers of interfering bacteria capable of growing on CT-SMAC.This work was supported with a grant awarded by the National Research Foundation of South Africa
Diversity and antimicrobial susceptibility profiling of staphylococci isolated from bovine mastitis cases and close human contacts
The objectives of this study were to examine the
diversity of Staphylococcus spp. recovered from bovine
intramammary infections and humans working in close
contact with the animals and to evaluate the susceptibility
of the staphylococcal isolates to different antimicrobials.
A total of 3,387 milk samples and 79 human
nasal swabs were collected from 13 sampling sites in
the KwaZulu-Natal province of South Africa. In total,
146 Staph. aureus isolates and 102 coagulase-negative
staphylococci (CNS) were recovered from clinical and
subclinical milk samples. Staphylococcus aureus was
isolated from 12 (15.2%) of the human nasal swabs and
95 representative CNS were recovered for further characterization.
The CNS were identified using multiplex-
PCR assays, matrix-assisted laser desorption/ionization
time-of-flight mass spectrometry (MALDI-TOF
MS), and tuf gene sequencing. Seven Staphylococcus
spp. were identified among the CNS of bovine origin,
with Staph. chromogenes (78.4%) predominating. The
predominant CNS species recovered from the human
nasal swabs was Staph. epidermidis (80%) followed by
Staph. chromogenes (6.3%). The antimicrobial susceptibility
of all staphylococcal isolates was evaluated using
disk diffusion and was supplemented by screening for
specific antimicrobial resistance genes. Ninety-eight
(67.1%) Staph. aureus isolates of bovine origin were
pansusceptible; 39 (26.7%) isolates were resistant to
a single class, and 7 (4.8%) isolates were resistant to
2 classes of antimicrobials. Two Staph. aureus (1.4%)
isolates were multidrug-resistant. Resistance to penicillin
was common, with 28.8% of the bovine and 75% of
the human Staph. aureus isolates exhibiting resistance.
A similar observation was made with the CNS, where
37.3% of the bovine and 89.5% of the human isolates
were resistant to penicillin. Multidrug-resistance was
common among the human CNS, with 39% of the
isolates exhibiting resistance to 3 or more classes of antimicrobials. The antimicrobial susceptibility results
suggest that resistance among staphylococci causing
bovine intramammary infections in South Africa is uncommon
and not a significant cause for concern. In contrast,
antimicrobial resistance was frequently observed
in staphylococcal isolates of human origin, highlighting
a possible reservoir of resistance genes. Continued
monitoring of staphylococcal isolates is warranted to
monitor changes in the susceptibility of isolates to different
classes of antimicrobials.University of Pretoria, RESCOM, the National Research Foundation (NRF) of
South Africa, and the KZN Department of Agriculture & Rural Development (South Africa).http://www.journals.elsevier.com/journal-of-dairy-science2016-09-30hb201
Identification and characterization of Staphylococcus devriesei isolates from bovine intramammary infections in KwaZulu-Natal, South Africa
BACKGROUND : Coagulase-negative staphylococci (CoNS) are among the leading bacterial causes of bovine mastitis
in many dairy-producing countries. Among the challenges associated with the specific diagnosis of CoNS infections
is the biochemical heterogeneity of the species in the genus and the unavailability of accurate, cost-effective and
up-to-date diagnostic tests. A previous study investigating the diversity of CoNS associated with cases of bovine
mastitis in South Africa, resulted in six CoNS isolates which could not be identified despite the use of a
combination of different molecular assays. The identification and characterisation of the isolates was pursued
further in this study.
RESULTS : The six CoNS isolates in question were identified by sequencing multiple housekeeping genes (dnaJ,
hsp60, rpoB, 16S rRNA) and characterized through the use of matrix-assisted laser/desorption ionization time of
flight mass spectrometry (MALDI-TOF MS) and the Biolog GEN III Microplateâ„¢ bacterial identification system.
Sequencing of housekeeping genes identified the isolates as S. devriesei. This Staphylococcus species was only
described in 2010 and this is the first report documenting the isolation of S. devriesei from cases of bovine IMIs in
South Africa. Analysis of mass spectra generated by the six isolates showed intra-species variation which was also
observed when evaluating the metabolic profiles of the isolates using the Biolog GEN III system. Neither the MALDITOF
MS nor the Biolog database are currently populated with data relating to S. devriesei, resulting in the isolates
not being identified, in the case of MALDI-TOF MS analysis, or mis-identified as was observed with the Biolog GEN
III system.
CONCLUSIONS : The phenotyping data collected during this investigation provides useful information concerning
Staphylococcus devriesei which could be used to populate user system databases thereby ensuring the accurate
identification of isolates in future. The availability of improved diagnostics will in turn facilitate studies to elucidate
the epidemiology, pathogenicity and true prevalence of this species in dairy herds.The research presented herein is funded, in part, by the University of
Pretoria, National Health Laboratory Services, RESCOM, the National Research
Foundation (NRF) Research Technology Fund and the KZN Department of
Agriculture and Rural Development. The MALDI-TOF MS work is supported in
part by the National Research Foundation (NRF) of South Africa (Grant
specific unique reference number, UID 74426).https://bmcvetres.biomedcentral.comMedical Microbiolog
Disruption in the Regulation of Immune Responses in the Placental Subtype of Preeclampsia
Preeclampsia is a pregnancy-specific disorder, of which one of its major subtypes, the placental subtype is considered a response to an ischemic placental environment, impacting fetal growth and pregnancy outcome. Inflammatory immune responses have been linked to metabolic and inflammatory disorders as well as reproductive failures. In healthy pregnancy, immune regulatory mechanisms prevent excessive systemic inflammation. However, in preeclampsia, the regulation of immune responses is disrupted as a result of aberrant activation of innate immune cells and imbalanced differentiation of T-helper cell subsets creating a cytotoxic environment in utero. Recognition events that facilitate immune interaction between maternal decidual T cells, NK cells, and cytotrophoblasts are considered an indirect cause of the incomplete remodeling of spiral arteries in preeclampsia. The mechanisms involved include the activation of immune cells and the subsequent secretion of cytokines and placental growth factors affecting trophoblast invasion, angiogenesis, and eventually placentation. In this review, we focus on the role of excessive systemic inflammation as the result of a dysregulated immune system in the development of preeclampsia. These include insufficient control of inflammation, failure of tolerance toward paternal antigens at the fetal–maternal interface, and subsequent over- or insufficient activation of immune mediators. It is also possible that external stimuli, such as bacterial endotoxin, may contribute to the excessive systemic inflammation in preeclampsia by stimulating the release of pro-inflammatory cytokines. In conclusion, a disrupted immune system might be a predisposing factor or result of placental oxidative stress or excessive inflammation in preeclampsia. Preeclampsia can thus be considered a hyperinflammatory state associated with defective regulation of the immune system proposed as a key element in the pathological events of the placental subtype of this disorder
Prevalence of carbapenem resistance genes in Acinetobacter baumannii isolated from clinical specimens obtained from an academic hospital in South Africa
Acinetobacter baumannii is an important cause of hospital-acquired infections. The occurrence of carbapenem resistance that is
caused by the carbapenem-hydrolysing class D β-lactamases and the metallo-β-lactamases (MBLs) limits the range of therapeutic
alternatives in treating A. baumannii infections. In this study, two multiplex polymerase chain reactions were performed to screen for both carbapenem-hydrolysing class D β-lactamases and MBL genes in 97 clinical isolates of A. baumannii. Oxacillinase (OXA)-51 had a prevalence of 83% (81/97), and OXA-23 had a prevalence of 59% (57/97). One isolate was positive for an MBL
[Verona integron-encoded metallo β-lactamases (VIM)]. Therefore, continuous surveillance and monitoring of A. baumannii is
crucial because of the high prevalence of antibiotic resistance genes.http://www.sajei.co.za/index.php/SAJEIam2013ay201
Prevalence of Clostridium difficile toxin genes in Pretoria
The hypervirulent polymerase chain reaction (PCR) ribotype 027 strain of Clostridium difficile produces toxins A, B and a binary
toxin. Toxin detection kits are commonly used in diagnostic laboratories, but have been unsuccessful in detecting all of the
relevant C. difficile strains, and the toxins produced. In this study, conventional PCR was used to detect the presence of the genes
of toxin A, toxin B and the binary toxin of C. difficile. Eighty-four frozen (collected between 2006-2007) and 13 fresh (collected in
2010) stool specimens, obtained in Pretoria, were analysed. The genes for toxin A, toxin B and the binary toxin were detected in
one of the fresh stool specimens. This may have implications for healthcare facilities, and suggests the possible emergence of
the highly virulent PCR ribotype 027 strain of C. difficile in Pretoria. This emphasises the importance of continuous surveillance
and monitoring of C. difficile outbreaks.http://www.sajei.co.za/index.php/SAJEIay201
Characteristics and outcomes of patients admitted to a tertiary academic hospital in Pretoria with HIV and severe pneumonia : a retrospective cohort study
BACKGROUND : Human immunodeficiency virus (HIV) contributes significantly to morbidity and mortality in South
Africa. Pneumonia and opportunistic infections remain a major cause for hospital admission among those living with
HIV, even in the era of the widespread availability of antiretroviral therapy.
METHODS : In this retrospective cohort study, the records of patients admitted with HIV and severe pneumonia,
requiring high care/intensive care admission, during a period of 12 months (February 2018 to January 2019) were
reviewed. Demographic details, antiretroviral use, HIV viral load, CD4 count, sputum culture results and radiological
imaging of patients were recorded. Data was analysed to determine variables associated with mortality.
RESULTS : One hundred and seventeen patient records were reviewed for this study. The patients were young (mean
age 38.3 years), had advanced disease with low CD4 counts (mean 120.2 cells/mm3) and high HIV viral loads (mean
594,973.7 copies/mL). Only 36.9% (42/117) were on highly active antiretroviral therapy (HAART) on presentation to the
hospital. Mycobacterium tuberculosis (M. tuberculosis) was found to be the cause for pneumonia in 35% (41/117), whilst
Pneumocystis jirovecii (P. jirovecii) was found in 21.4% (25/117). Bacterial pneumonia was the cause in 17.1% (20/117)
of patients while no specific aetiology was found in 26.6% (31/117) of patients in the cohort. Mortality among the
cohort studied was high (40.1%) and the average length of stay in hospital in excess of two weeks. The need for ICU
admission, ventilation and CMV viremia was associated with increased mortality. Chest X-ray findings did not correlate
with the aetiology of pneumonia, but multiple B-lines on lung ultrasound correlated with P. jirovecii as an aetiology
and there was a signal that pleural effusion with fibrin stranding predicts tuberculosis.
CONCLUSIONS : Patients studied presented with advanced HIV and were often naïve to antiretroviral therapy. Mortality
in this cohort of young patients was high, which emphasis the need for earlier diagnosis and treatment of HIV at a
primary care level. Lung ultrasound may have clinical utility in the management of patients with HIV and pneumonia,
particularly to diagnose P. jirovecii as an aetiology.http://www.biomedcentral.com/bmcinfectdisam2023Internal MedicineMedical Microbiolog
High prevalence of oxacillinases in clinical multidrug-resistant Acinetobacterbaumannii isolates from the Tshwane region, South Africa - an update
BACKGROUND : Acinetobacter baumannii is an important hospital-acquired pathogen in healthcare facilities that
frequently causes bacteraemia and ventilator-associated pneumonia in intensive care units. Acinetobacter baumannii
can be isolated from various sites in the hospital environment like medical equipment, bed linen, medical personnel
and indwelling catheters. It is difficult to treat A. baumannii infections because of their highly resistant antimicrobial
profiles. The purpose of this study was to determine the prevalence of β-lactamase genes in multidrug-resistant (MDR)
clinical A. baumannii isolates using Multiplex-PCR (M-PCR) assays.
METHODS : One hundred MDR A. baumannii isolates were collected from the diagnostic division of the Department of
Medical Microbiology after routine analysis of the submitted specimens. All collected isolates were identified and tested
for susceptibility using the VITEK 2® system (bioMérieux, France). Six isolates were excluded from this study because the
isolates were incorrectly identified as A. baumannii with the VITEK 2® system (bioMérieux, France). Molecular tests, namely
M-PCR assays, pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) were performed. MLST
analyses were performed on representative isolates from the four major pulsotypes (≥5 isolates with 80 % similarity) and
selective isolates from each minor pulsotype.
RESULTS : All the A. baumannii isolates showed 100 % resistance to ampicillin, amoxicillin, cefuroxime, cefuroximine axetil,
cefoxitin, cefotaxime and nitrofurantoin. Seven percent of the isolates were resistant to amikacin. Two percent of the
isolates were classified as having intermediate susceptibility to tigecycline. A. baumannii isolates showed an antibiotic
resistance profile of 67 % and higher to antibiotics, such as ceftazidime, cefepime, imipenem, meropenem, gentamicin,
ciprofloxacin and trimethoprim/sulfamethoxazole. None of the isolates were resistant to colistin. The M-PCR assays
showed that 99 % of the isolates contained the OXA-51 gene and 77 % contained the OXA-23 gene. None of the
isolates contained the GES, GIM, IMP, KPC, NDM, OXA-24, OXA-58, PER, SIM, SPM, VEB and VIM genes. Representative A.
baumannii isolates were grouped into five existing sequence types (ST): ST106, ST258, ST339, ST502, ST758 and ST848.
Isolates belonging to the pan-European clonal lineages I and II (EUI and EUII) were identified.
CONCLUSION : The high prevalence of MDR A. baumannii isolates has a severe impact on available treatment choices and
this in return impacts on treatment outcomes in the studied healthcare facilities. The most dominant ST among the
collected isolates was ST758, member of the EUI group. The presence of the OXA-23 gene was not restricted to a
specific ST. Continuous research and surveillance is necessary to monitor the circulating β-lactamase genes in clinical
settings to guide infection control policies in order to try and curb the spread of this bacterium.ML was supported by a
National Research Foundation (NRF) grant. The MALDI-TOF analysis is based on
research supported in part by the National Research Foundation (NRF) of South
Africa (Grant specific unique reference number (UID) 74426).http://www.biomedcentral.com/bmcinfectdis/am201
Normal flora and bacterial vaginosis in pregnancy : an overview
The female genital tract is an intricate, yet balanced ecosystem that hosts a variety of
different residential microflora. The physiological changes that occur during pregnancy may
disrupt this balanced ecosystem and predispose women to a potentially pathogenic
microbiota. Bacteria that are associated with bacterial vaginosis (BV) are opportunistic
pathogens that frequently form part of this microbiota. The overgrowth of and infections
with these bacteria are linked to poor obstetric outcomes and increased transmission of other
reproductive tract infections (RTIs). These infections increase women’s susceptibility of
acquiring HIV, the rates of HIV shedding and the development of Acquired Immune Deficiency Syndrome (AIDS) in HIV infected patients. It is unknown how the plethora of
bacterial species associated with BV contributes to the dynamics of this condition. The use
of high-throughput methods have led to the in-depth investigation of different BV-related
bacterial species and the functional capabilities of these species. However, the pathogenesis
of BV is still poorly defined and the role of individual BV-related bacterial species in specific
pregnancy complications is unclear and controversial. The majority of BV infections are
asymptomatic and successful diagnosis is complicated by the lack of reliable and
standardized diagnostic tests.University of Pretoria, the Medical Research Council
(South Africa) and the National Health Laboratory Service (NHLS).http://www.tandfonline.com/loi/imby202017-05-31hb2016Medical Microbiolog
Assessment of Atopobium vaginae and Gardnerella vaginalis concentrations in a cohort of pregnant South African women
OBJECTIVES : The purpose of this cross-sectional study was to assess Atopobium vaginae and Gardnerella vaginalis concentrations in pregnant women of different age groups, gestational age groups, vaginal flora categories and HIV status, and also to determine which DNA concentrations best discriminated between bacterial vaginosis (BV)-positive and non-BV categories. METHODS : Self-collected vaginal swabs were obtained from 220 pregnant women attending an antenatal clinic in Pretoria, Gauteng, South Africa, from July 2012 to December 2012. BV was detected with the Nugent scoring system, and A. vaginae and G. vaginalis DNA was quantified with a multiplex quantitative real-time PCR assay. RESULTS : Median concentrations of A. vaginae and G. vaginalis were not significantly different among various age groups (A. vaginae p=0.98 and G. vaginalis p=0.18) or different trimesters (A. vaginae p=0.31 and G. vaginalis p=0.19), but differed significantly among the vaginal flora categories (A. vaginae p<0.001 and G. vaginalis p<0.001) and HIV status (A. vaginae p<0.001 and G. vaginalis p=0.004). The presence of A. vaginae (OR=5.8; 95% CI 1.34 to 25.21 and p value=0.02) but not that of G. vaginalis (OR=1.90; 95% CI 0.81 to 4.43 and p value=0.14) was associated with HIV infection. An A. vaginae DNA concentration of ≥107 copies/mL together with a positive G. vaginalis result (≥100 copies/mL) best discriminated between BV-positive (39/220) and non-BV categories (181/220) with a sensitivity of 85% (95% CI 0.70 to 0.94) and a specificity of 82% (95% CI 0.76 to 0.88). CONCLUSION : A. vaginae and G. vaginalis were present in high numbers and concentrations in this pregnant cohort. Threshold concentrations should be established for specific populations to ensure sensitive molecular assays for BV detection.The University of Pretoria and the Medical Research Council (South Africa).http://sti.bmj.com2018-09-30hj2017Medical Microbiolog
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