Antimicrobial profile of coagulase-negative staphylococcus isolates from categories of individuals at a neonatal intensive care unit of a tertiary hospital, Ghana.

Introduction: we compared the antimicrobial resistance profile of young infants' clinical isolates (from blood samples) of Staphylococcus epidermidis and haemolyticus with those colonizing mothers, clinical staff, and students. Also, screened for resistance to the watch and reserve classified groups, antibiotics not prescribed in the Ho Teaching Hospital (HTH), Ghana. Methods: a cross-sectional study was conducted from March to June 2018 to determine the antimicrobial susceptibility of twenty-one antimicrobials for 123 isolates consisting of 54 S. epidermidis and 69 S. haemolyticus cultured from the participants. VITEK 2 was used for antimicrobial susceptibility testing. Staphylococcal species were identified using matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF). Statistical analysis was done with Grad-Pad prism. Results: for S. epidermidis, clinical staff isolates have the highest methicillin-resistant (65%), followed by young infants' (50%) and mothers' and students' twenty-five percent each. Both young infants and clinical staff's Staphylococcus haemolyticus isolates have 100% methicillin-resistant, while mothers' and students' ones have 82% and 63%, respectively. We have identified resistance to one watch (teicoplanin), two reserves (tigecycline and fosfomycin) antimicrobial groups, and mupirocin, an unclassified group. Conclusion: identifying coagulase-negative staphylococci (CoNS) resistance to watch and reserve groups of antimicrobials in a non-previously exposed hospital calls for further studies to determine molecular mechanisms of resistance to these antimicrobials.

The influence of unconventional ultrasonic pasteurization on the characteristics of curds obtained from goat milk with the low cholesterol content.

This study aimed to evaluate the influence of different power-time ultrasound regimes of pasteurization on the physical, chemical, organoleptic properties, and lipid quality indices of goat curds characterized by a low cholesterol level. Cholesterol was eliminated by a percentage of 92.1 % by treating the raw goat milk with beta-cyclodextrin in the proportion of 0.6 %. Afterward, the goat milk was subjected to the following ultrasound regimes: 320 W for 1 (PA1), 3 (PA3), and 6 min (PA6) and 881 W for 1 (PP1), 3 (PP3), and 6 min (PP6) and then used for the curds production. Due to the ultrasound treatment, the milk suffered a concentration phenomenon, the most accentuated being registered for the PP6 sample. Considering the sensory properties, the most appreciated curd was the one obtained by the PP6 regime which recorded the highest scores for color and taste. Regarding the microbiological aspects, the ability of ultrasounds to inactivate microorganisms is observed and the most accentuated phenomenon is reported in the PP6 case. Thus, in comparison with the control sample, the total number of germs is reduced by a proportion of 91.85 %, the ß-glucuronidase-positive Escherichia coli decreased by 93.15 %, while the coagulase-positive staphylococci were completely inactivated for the PP6 curd. The curds obtained for the PA6 and PP6 regimes registered the highest dry matter values as a cause of an accentuated syneresis process. The acidity values were higher for the curds obtained for PA1, PA3, and PA6 regimens due to more pronounced lactose hydrolysis and lower in the cases of PP3 and PP6 regimens compared to the control cheese. Twenty-five saturated, monounsaturated, and polyunsaturated fatty acids were identified in the curd samples and a rise in the unsaturated fatty acids proportion as the intensity of the applied ultrasound regime increased was observed. Also, AI, TI, and H/H lipid quality indices recorded better values as the power and time of the ultrasound action increased.

Anti-staphylococcal lysin, LSVT-1701, activity: In vitro susceptibility of Staphylococcus aureus and coagulase-negative staphylococci (CoNS) clinical isolates from around the world collected from 2002 to 2019.

LSVT-1701 (previously known as SAL200), is a novel, recombinantly-produced, bacteriophage-encoded lysin that specifically targets staphylococci via cell wall enzymatic hydrolysis. In vitro activities of LSVT-1701 and comparators were tested against 415 Staphylococcus aureus and coagulase-negative staphylococci (CoNS) clinical isolates expressing various resistance phenotypes. The isolates were collected worldwide from 2002 to 2019 and tested for in vitro susceptibility using broth microdilution methodology performed according to the Clinical and Laboratory Standards Institute (CLSI) guidelines. Minimum inhibitory concentration (MIC) interpretations were based on CLSI and EUCAST criteria. MIC90 for all S. aureus, methicillin-susceptible S. aureus, methicillin-resistant S. aureus, and CoNS, were 2, 2, 2 and 2 µg/ml, respectively. LSVT-1701's activity was not adversely affected by resistance to antimicrobial comparators against this worldwide collection of S. aureus and CoNS clinical isolates. The results of this study support further clinical development of LSVT-1701 to treat staphylococcal infections, including those caused by multidrug resistance isolates.

Characterization of biofilms and antimicrobial resistance of coagulase-negative Staphylococcus species involved with subclinical mastitis.

Biofilm formation is a central feature to guarantee staphylococcal persistence in hosts and is associated with several diseases that are difficult to treat. In this research paper, biofilm formation and antimicrobial susceptibility were investigated in staphylococcal strains belonging to several species. These strains were isolated from the milk of cows with subclinical mastitis and most of them were coagulase-negative, with the prevalence of Staphylococcus chromogenes. High genetic diversity was observed among the strains by pulsed field gel electrophoresis. Antimicrobial resistance was assessed by disk diffusion and more than 50% of the strains were resistant to ampicillin and penicillin G, with multi-resistance profiles (13.6%) also being observed. Most strains (65.9%) formed biofilms when cultivated in BHI supplemented with 1% glucose. Most strains (72.7%) carried the intercellular adhesion gene (icaA), while less than half (36.3%) carried the biofilm-associated protein gene (bap). Concentrations of up to 10xMIC of erythromycin and tetracycline were not sufficient to suppress cell viability in preformed biofilms. Our results revealed that a genetically diverse group of biofilm-forming Staphylococcus species can be involved in subclinical mastitis. Since high antimicrobial concentrations cannot eradicate biofilm cells in vitro, their use in dairy animals may be ineffective in controlling infections, while supporting selection of resistant microorganisms. These data reinforce the need for alternative therapies aiming at disrupting biofilms for effective disease control.

Coagulase negative Staphylococcus bacteremia in hematopoietic stem cell transplant recipients: Clinical features and molecular characterization.

The purpose of our study was to investigate the epidemiology of coagulase negative staphylococci (CoNS) responsible for bacteremia in hematopoietic stem cell transplant (HSCT) recipients and to determine the prevalence and the genetic background of methicillin resistance. The prevalence of CoNS bacteremia was 7.4% (54/728), higher in allograft (10.7%) than in autograft (4.7%) recipients. A sepsis or a septic shock were observed in 9% of cases. No deaths were attributable to CoNS bacteremia. The methicillin resistance rate was 81%. All MR-CoNS, harbored mecA gene and 90% were typeable with SCCmec typing using PCR amplification. The SCCmec type IV was the most frequent (44%). Clonal dissemination of MR- Staphylococcus epidermidis strains was limited. Our study showed a low prevalence and favorable outcome of CoNS bacteremia in HSCT recipients with limited clonal diffusion. However, they were associated with a significant rate of severe infections and a high rate of methicillin resistance, mediated by SCCmec IV element in most cases.

Microbiological evaluation of pasty and soft ice cream / Avaliação microbiológica de sorvetes pastosos e expressos / Evaluación microbiológica de helados pastosos y suaves

Ice cream is susceptible to contamination by handling and bad hygiene conditions during both the storage process and the fractioning for sale, and once contaminated, it can cause diseases. The purpose of this survey was to evaluate the microbiological quality of ice cream sold in bulk, of pasty and soft types, offered for consuming. Thirty samples of pasty ice cream sold in bulk, and thirty samples of soft ice cream were analyzed through the counting of thermotolerant coliforms, coagulase-positive Staphylococcus spp., and searching for the presence of Salmonella spp. During the study, a total of ten (33%) samples of pasty ice cream and five (16%) samples of soft ice cream were found to be beyond the limits established by the Brazilian law. Salmonella spp. was found in four samples (6.7%). These results are an alert for the need of greater attention to the microbiological quality of ice cream in order to ensure the safety of its consumers.(AU)

Os sorvetes são suscetíveis à contaminação pela manipulação e más condições higiênicas durante o processamento, armazenamento e do fracionamento para venda, uma vez contaminados podem causar doenças. O objetivo deste estudo foi avaliar a qualidade microbiológica de sorvetes, vendidos a granel, pastosos e expressos, oferecidos para consumo. Trinta amostras de sorvete pastoso, vendido a granel, e trinta amostras de sorvete expresso foram analisadas realizando-se contagem de coliformes termotolerantes, Staphylococcus spp. coagulase-positiva e pesquisando-se a presença de Salmonella spp. Foram detectadas dez (33%) amostras de sorvete pastoso e cinco (16%) amostras de sorvete expresso fora dos limites estabelecidos pela legislação brasileira. Salmonella spp. foi encontrado em quatro amostras (6,7%). Esses resultados alertam para a necessidade de uma maior atenção à qualidade microbiológica dos sorvetes, a fim de garantir a segurança do consumidor.(AU)

Los helados son susceptibles a la contaminación por manipulación y malas condiciones higiénicas durante el procesamiento, almacenamiento y fraccionamiento para venta, una vez contaminados pueden causar enfermedades. El objetivo de este estudio ha sido evaluar la calidad microbiológica de helados vendidos a granel, pastosos y suaves, ofrecidos para el consumo. Se analizaron treinta muestras de helados pastosos vendidos a granel, y treinta muestras de helados suaves, realizándose el conteo de coliformes termotolerantes, Staphylococcus spp. coagulase positiva e investigándose la presencia de Salmonella spp. Se detectaron diez (33%) muestras de helado pastoso y cinco (16%) muestras de helado blando fuera de los límites establecidos por la legislación brasileña. Salmonella spp. se encontró en cuatro muestras (6,7%). Esos resultados destacan la necesidad de una mayor atención a la calidad microbiológica de los helados, con el fin de garantizar la seguridad del consumidor.(AU)

Antibacterial and anti-biofilm activities of walnut pellicle extract (Juglans regia L.) against coagulase-negative staphylococci.

Juglans regia L. (common walnut) is a deciduous tree belonging to Juglandaceae family. Since ancient time, walnut was widely used in traditional medicine for its antioxidant, antidiabetic, antimicrobial, anti-inflammatory, anti-atherogenic and liver-protective effects. In this work, the antibacterial and anti-biofilm activities of walnuts pellicle extract against coagulase-negative staphylococci were evaluated. Qualitative chemical analysis was performed by the thin layer chromatography. UPLC-Ms/Ms was used to identify the chemical composition of J. regia extract. The total flavonoid and phenolic contents were determined by the Aluminium chloride and Folin-Ciocalteu methods, respectively. The extract showed antibacterial activity with MIC ranging from 3.60 to 461.75 µg/ml and MBC ranging from 461.75 to >461.75 µg/ml. Furthermore, it significantly reduced biofilm biomass and cell viability in a dose-dependent manner. Biological activities of J. regia extract may be due to its high flavonoid and phenolic contents. The obtained results are promising and they deserve further scientific investigations.

Clinical characteristics and relevance of coagulase-negative Staphylococci other than S. epidermidis by positive blood culture.

INTRODUCTION: Staphylococci is the most commonly isolated bacteria in blood cultures (BC), and is mostly regarded as contamination. However, the clinical relevance and characteristics of CNS other than Staphylococcus epidermidis (Non-SE-CNS) is still unknown. METHODS: For the purpose of clarifying the epidemiology and clinical significance of Non-Se-CNS infections, we retrospectively evaluated BC isolates of Non-Se-CNS at our institute from May 2013 to March 2017. Infections were defined as true bacteremia if (1) two or more positive BCs of the same species were present (or detection of the same species in another clinically relevant sample), (2) the patient had clinical symptoms as outlined in the Centers for Disease Control and Prevention's definition of primary blood stream infection8 and the symptoms or markers of inflammation (e.g., fever, symptoms, white cell counts, C-reactive protein, or procalcitonin) were improved after pathogen-directed therapy (e.g. either antibiotic therapy and/or removal of the foreign body in the case of catheter-related blood stream infections suspected) and (3) without any other infection. RESULTS: During the period, 279 patients with BC positive for Non Se-CNS were identified. 44 patients were excluded, either due to missing data or multiple pathogens in the same BC. Among the remaining 235 patients, 67 (29%) were categorized as true bacteremia (n = 43) or possible infections (n = 24). Resistance rate to methicillin among Non-SE-CNS is about 60%, which is lower than those among S. epidermidis of 78%. CONCLUSION: About 30% of Non-Se-CNS isolates were clinically relevant in our study. Antibiotic susceptibility of Non-SE-CNS is quite different than those of S. epidermidis.

Short communication: Characterization of an atypical maltose-negative Staphylococcus aureus through the use of phenotypic and molecular techniques.

The most clinically relevant staphylococci in veterinary medicine are those that are coagulase-positive, namely Staphylococcus aureus. During microbiological udder health monitoring (2009-2018), a new S. aureus strain (coagulase-positive and maltose-negative) was discovered as an emerging udder pathogen during routine examinations of South African dairy herds. This study challenged the conventional microbiological diagnosis of staphylococci by comparing its results to those of the MALDI-TOF mass spectrometry and 16S rRNA sequencing. Both of these tests confirmed that the maltose-negative staphylococcus (MNS), identified as Staphylococcus pseudintermedius by conventional microbiology, was S. aureus ST2992. Multi locus sequence typing was performed on 3 of the MNS isolates and indicated that these isolates were of single origin. These strains tested positive for both MALA and MALR genes (control: S. aureus ATCC 25923). Although the α-glucosidase gene was present, it was not expressed phenotypically. The latter may be attributed to the abnormal stop codon identified in the MALA gene sequence of S. aureus ST2992 (GenBank accession number, MN531305). The newly identified MNS has a field behavior different to that of maltose-positive S. aureus, and more similar to the low virulence of non-aureus staphylococci.

Detection of sasX Gene and Distribution of SCCmec Types in Invasive and Non-invasive Coagulase-negative Staphylococci

Background: Coagulase-negative staphylococci, which belong to the normal microbiota of the skin and mucous membranes, are opportunistic pathogens. sasX, a newly described protein, is thought to play an important role in nasal colonization and methicillin-resistant Staphylococcus aureus virulence, and it may be acquired from coagulase-negative staphylococci by horizontal gene transfer. It has been considered that understanding the function of sasX gene may help clarify the relevance of the different adhesion mechanisms in the pathogenesis of infections associated with biofilm. Aims: To investigate the sasX gene presence, staphylococcal cassette chromosome mec types, and antimicrobial resistance patterns of invasive and noninvasive coagulase-negative staphylococci isolates. Study Design: Cross-sectional study. Methods: The study included a total of 180 coagulase-negative staphylococci strains. Non-invasive isolates (n=91) were obtained from the hands of healthy volunteers who do not work at the hospital (n=30), the nasal vestibule of healthy volunteer hospital workers (n=26), and central venous catheter (n=35). Invasive isolates (n=89) were isolated from peripheral blood cultures of inpatients who do not have catheters. All isolates were identified by conventional microbiological methods, automated systems, and, if needed, with matrix-assisted laser desorption/ionization-time of flight. Staphylococcal cassette chromosome mec typing, sasX and mec gene detection, antibiotic susceptibility, and sasX gene sequence analysis were performed. Results: Peripheral blood, central venous catheter colonization, and nasal vestibule isolates were positive for the sasX gene, whereas hand isolates were negative. sasX gene was present in 17 isolates, and no statistical significance was found between invasive and noninvasive isolates (p=0.173). Sequence analysis of the sasX genes showed high homology to related proteins of Staphylococcus phage SPbeta-like and Staphylococcus epidermidis RP62A. staphylococcal cassette chromosome mec type V was the most prevalent regardless of species. staphylococcal cassette chromosome mec type II was more frequent in invasive isolates and found to be statistically important for invasive and noninvasive S. epidermidis isolates (p=0.029). Staphylococcus haemolyticus isolates had the overall highest resistance rates. Resistance to ciprofloxacin, trimethoprim-sulfamethoxazole, and erythromycin was found to be higher in isolates from catheter and blood culture. Staphylococcus hominis isolates had the highest rate for inducible clindamycin resistance. None of the isolates were resistant to vancomycin, teicoplanin, and linezolid. Conclusion: The sasX gene is detected in 9.44% of the isolates. There is no statistical difference between the sasX-positive and -negative isolates in terms of antibacterial resistance and the presence of sasX and SCCmec types. Further studies about the role of sasX at virulence in coagulase-negative staphylococci, especially from clinical samples such as tracheal aspirate and abscess isolates, and distribution of staphylococcal cassette chromosome mec types are needed.

Detection of bacteria in bovine ovarian follicular fluid.

The aim of this study was to examine the sterility of follicular fluid in large ovarian follicles in dairy cows. In all, 17 samples of paired follicular fluids and uterine contents collected from post-slaughtered dairy cows were cultured to detect aerobic and facultative anaerobic bacteria. Furthermore, the origin of the bacterial isolates from samples of follicular fluid and the uteri was also investigated using PFGE analysis. Follicular fluid concentrations of lipopolysaccharides were also determined. Of 17 uterine samples, 15 (88%) were detected as contaminated. In total, nine different bacterial genera and species were identified in the uterine and follicular fluid samples. Escherichia coli was the most prevalent bacterial species isolated from the uterine samples. Out of seven isolates of Staphylococcus aureus from the uterine samples, 6 (85%) were coagulase positive. Six isolates of Staphylococcus spp. were identified in 6 out of 17 follicular fluid samples (35%). Two out of six isolates were identified as Staphylococcus aureus (33%). Our results show that ovarian follicular fluid is not sterile in the bovine. The presence of Staphylococcus aureus in follicular fluid may partly explain the occurrence of infertility in some dairy cows. SIGNIFICANCE AND IMPACT OF THE STUDY: The results of the present study show that ovarian follicular fluid is not sterile in bovines. The presence of Staphylococcus aureus in follicular fluid may partly explain the occurrence of infertility in some dairy cows.

Antimicrobial resistance patterns and biofilm formation of coagulase-negative Staphylococcus species isolated from subclinical mastitis cow milk samples submitted to the Onderstepoort Milk Laboratory.

BACKGROUND: Increased prevalence of antimicrobial resistance, treatment failure, and financial losses have been reported in dairy cows with coagulase-negative Staphylococcus (CoNS) clinical mastitis, however, studies on CoNS infections are limited in South Africa. Therefore, the objectives of this study were to investigate the antimicrobial resistance patterns and biofilm formation in CoNS isolated from cow milk samples submitted to the Onderstepoort Milk Laboratory. RESULTS: A total of 142 confirmed CoNS isolates were used for this study. Biofilm formation was identified in 18% of CoNS tested. Staphylococcus chromogenes (11%) had the highest proportion of biofilm formation followed by S. haemolyticus (4.0%), S. epidermidis, S. hominis, S. xylosus, and S. simulans with 1% respectively. Ninety percent (90%) of CoNS were resistant to at least one antimicrobial (AMR) and 51% were multidrug-resistant (MDR). Resistance among CoNS was the highest to ampicillin (90%) and penicillin (89%), few isolates resistant to cefoxitin and vancomycin, 9% respectively. Similarly, MDR-S. haemolyticus (44%), MDR-S. epidermidis (65%), and MDR-S. chromogenes (52%) were mainly resistant to penicillins. The most common resistance patterns observed were resistance to penicillin-ampicillin (16%) and penicillin-ampicillin-erythromycin (10%). Only 42% of biofilm positive CoNS were MDR. CONCLUSION: The majority of CoNS in this study were resistance to penicillins. In addition, most isolates were ß-lactam resistant and MDR. Biofilm formation among the CoNS in this study was uncommon and there was no significant difference in the proportion of MDR-CoNS based on the ability to form a biofilm.

Eradication of Repeated Episodes of Coagulase-Negative Staphylococcus Peritonitis: a Multipronged Approach.

Bacterial peritonitis continues to be a significant cause of morbidity and mortality in patients undergoing peritoneal dialysis. Episodes of peritonitis due to coagulase-negative staphylococci (CNS) are especially prone to relapse or repeat, often culminating in catheter removal and technique failure. This pattern is thought to be associated with the production of a biofilm that offers protection against antimicrobials and the host defense systems. We report the successful eradication of repeated episodes of CNS peritonitis using an aggressive multipronged therapeutic approach combining several antibiotics and a thrombolytic agent.

Characterization of virulence factors, antifungal resistance with ERG-11 gene among Candida species isolated from pulmonary samples.

BACKGROUND: Candida is a part of the normal oropharyngeal flora and the upper respiratory tract. Candida albicans(C. albicans), is the predominant species causing respiratory tract infections associated with pneumonia. Resistance to azole antifungal agents among the C. albicans may be due to alteration of the target enzymes, which are encoded in ERG11 gene. The biofilm formation may also be a cause to antifungal resistance. MATERIALS AND METHODS: This study was conducted at Chettinad Hospital and Research Institute. Samples were collected from June 2018-June 2019, for a period of 1 year. After species confirmation, virulence factor among the Candida species were identified by hemolysis test, coagulase test and biofilm formation. Genotypic confirmation of C. albicans and their azole resistance due to ERG 11 gene were done using multiplex PCR. RESULTS: In our study, 31 (55%) C. albicans, 8 (14%) Candida glabrata(C. glabrata) and 10 (17%) Candida tropicalis(C. tropicalis), three Aspergillus flavus(A. flavus), two Aspergillus fumigatous (A. fumigatous), one Aspergillus niger (A. niger) and one Mucor species were isolated. In C. albicans, 31 were positive for Germ tube and Chalmydospore formation. Six of candida species were isolated along with bacterial co infection. Among the Candida isolates, 17 (55%) C. albicans strains were strongly biofilm positive and 14(45%) were negative. The susceptibility pattern of (n = 31) C. albicans were as follows: fluconazole (21(68%) S, 10(32%) R), voriconazole (22(71%)S),9(21%) R) and Amphotericin B 31(100%) S). Among the 19 C. albicans, four were positive for ERG11 gene. CONCLUSION: The isolation of C. albicans and non - albicans from respiratory specimens should be reconsidered as these organisms are re-emerging pathogens. Speciation is needed due to variation in species pathogenicity and their susceptibility.

Transfer of a lincomycin-resistant plasmid between coagulase-negative staphylococci during soybean fermentation and mouse intestine passage.

Staphylococcus equorum is a benign bacterium and the predominant species in high-salt fermented food. Some strains of S. equorum contain antibiotic-resistance plasmids, such as pSELNU1 that contains a lincosamide nucleotidyltransferase (lnuA) gene and confers resistance to lincomycin. Previously, we showed that pSELNU1 is transferred to other bacteria under laboratory growth conditions. However, it is not known if the plasmid can be transferred to other bacteria during food fermentation (in situ) or during passage through animal intestines (in vivo). In this study, we examined the in situ and in vivo transfer of pSELNU1 using Staphylococcus saprophyticus as a recipient. During soybean fermentation, pSELNU1 was transferred to S. saprophyticus at a rate of 1.9 × 10-5-5.6 × 10-6 per recipient in the presence of lincomycin. However, during passage through murine intestines, the plasmid was transferred at similar rates (1.3 × 10-5 per recipient) in the absence of lincomycin, indicating that the plasmid transfer is much more efficient under in vivo conditions. Based on these results, we conclude that it is prudent to examine food fermentation starter candidates for the presence of mobile genetic elements containing antibiotic resistance genes and to select candidates lacking these genes.

The species distribution, antimicrobial resistance and risk factors for poor outcome of coagulase-negative staphylococci bacteraemia in China.

Objective: Coagulase-negative staphylococci (CoNS) are one of the major opportunistic pathogens and the incidence of CoNS bacteraemia is increasing. However, most of the CoNS-positive blood cultures are contaminants rather than true CoNS bacteraemia. In order to minimize contamination, we defined true CoNS bacteraemia as the patient that has two or more identical CoNS-positive blood cultures drawn within 48 h in this study and the objective of this study was to analyse the species distribution and antibiotic resistance and to identify risk factors for 30-day mortality of the true CoNS-bacteraemia. Method: By reviewing the electronic medical database, this study retrospectively analysed patients diagnosed as CoNS bacteraemia by blood cultures in a comprehensive tertiary care hospital in China from January 1, 2014, to December 31, 2017. Result: A total of 1241 patients with 1562 episodes of CoNS-positive blood cultures were recorded in the database but only 157 patients were finally diagnosed as true CoNS bacteraemia after contaminants were excluded. All these 157 patients (12.7%, 157/1241) had bacteraemia-related clinical symptoms. Among the 157 patients, the most common species were Staphylococcus hominis (40.8%), Staphylococcus epidermidis (36.3%) and Staphylococcus capitis (11.5%). The antimicrobial susceptibility tests showed that all CoNS had a high rate of resistance to penicillin (94.9%), oxacillin (93.6%) and erythromycin (92.4%). Resistance to gentamicin (22.3%) and rifampicin (10.8%) was low, and none of the bacteria were resistant to vancomycin or linezolid. The 30-day mortality of patients with CoNS bacteraemia was up to 12.7% (20/157), and the multivariate logistics regression analysis showed that chronic renal failure (OR 5.9, 95% CI 1.6-21.5, p = 0.007) and chronic liver failure (OR 4.0, 95% CI 1.2-13.1, p = 0.024) were both the significant independent risk factors for the 30-day mortality of CoNS bacteraemia. Conclusion: Staphylococcus hominis and Staphylococcus epidermidis were the most common species in CoNS bacteraemia. All CoNS had high multi-drug resistance, but gentamicin and rifampicin had a relatively lower resistance and could be considered as alternative antibiotics for anti-CoNS bacteraemia in addition to vancomycin and linezolid. Additionally, patients with chronic renal failure or chronic liver failure have a higher 30-day mortality after the onset of CoNS bacteraemia.

Evaluation of Antibiotic Susceptibility Pattern and Distribution of coa Genes in Coagulase-negative S. aureus from Ahvaz, Iran.

BACKGROUND: Staphylococcus aureus is one of the important causes of clinical infections that can be more destructive by its antibiotic resistant strains. OBJECTIVE: This study aimed to determine the antibiotic susceptibility pattern and distribution of mecA and coa genes in clinical isolates of S. aureus. METHODS: Two hundred seventy-three specimens suspected to S. aureus were taken from hospitals of Ahvaz, southwest of Iran. Isolates were identified by standard microbiologic tests and confirmed by the molecular method. Antimicrobial susceptibility testing was carried out by disk diffusion method. The presence of mecA and coa genes was determined by PCR method. RESULTS: Of a total of 200 isolates which were tested for coagulase tube test, 143 (71.5%) showed coagulase positive, and 57 (28.5%) showed a coagulase-negative reaction. Antibacterial susceptibility pattern of 200 S. aureus isolates showed the highest and lowest susceptibility rate to linezolid (98%) and ciprofloxacin (42%), respectively. The prevalence of methicillin-resistant S. aureus (MRSA) by detection of mecA gene was estimated as 47.5 % (95/200), of which the rate of MRSA in coagulase positive and negative isolates was 35% (50/143), and 65% (45/57), respectively. Meanwhile, coa gene was detected in 100% of coagulase positive and 28.1% of coagulasenegative isolates. CONCLUSION: The results of this study showed that the number of atypical CNSA in our area is high. Since the coagulase test is an essential test for diagnosis of S. aureus, our findings regarding the emergence of CNSA are a warning about the misdiagnosis and selection of appropriate treatment approach for S. aureus isolates.

Acute-phase inter-alpha-trypsin inhibitor heavy chain 4 (ITIH4) levels in serum and milk of cows with subclinical mastitis caused by Streptococcus species and coagulase-negative Staphylococcus species.

The aim of the study was to investigate the concentrations of acute-phase inter-α-trypsin inhibitor heavy chain 4 (ITIH4) in serum and milk of cows with subclinical mastitis caused by Streptococcus spp. (STR) and coagulase-negative Staphylococcus spp. (CNS) and healthy cows. The blood and milk samples were obtained from 60 mid-lactation, multiparous Holstein-Friesian cows from 7 herds in the Lublin region of Poland. In the milk samples from 40 cows with subclinical mastitis, Streptococcus spp. and CNS were isolated. The ITIH4 was significantly higher in serum of cows with subclinical mastitis caused both by STR and CNS compared with healthy cows. One hundred percent of animals infected with Streptococcus spp. and 89% of animals infected with Staphylococcus spp. showed ITIH4 concentration in sera higher than 0.5 mg/mL. The concentration of ITIH4 in milk also was significantly higher in cows with subclinical mastitis caused by Streptococcus spp. and Staphylococcus spp. compared with the control group. Seventy percent of cows infected by STR and CNS showed ITIH4 concentration in milk higher than 2.5 µg/mL. Milk ITIH4 concentration higher than 5 µg/mL was found in 55% of animals infected with Streptococcus spp. and in 40% of animals infected with Staphylococcus spp. No statistically significant differences were observed in ITIH4 concentrations both in serum and in milk between the studied unhealthy animal groups. These results suggest that ITIH4 may be used in the future as a novel diagnostic marker in serum and in milk of subclinical mastitis in cows.

Short communication: Characterization of enterotoxin-producing Staphylococcus aureus isolated from mastitic cows.

Staphylococcus aureus is not only a common cause of bovine mastitis, but also an agent of food poisoning in humans. In an attempt to determine whether staphylococci causing bovine mastitis could also cause food poisoning, 60 isolates of presumed S. aureus were isolated in the period between March and August 2017 from 3,384 routine, composite, quarter milk samples of individual cows raised on 12 dairy farms in central Italy. Seventeen out of 60 isolates were confirmed as S. aureus after coagulase, thermonuclease, and biochemical tests. These isolates were analyzed by PCR for the presence of the nuc, sea, seb, sec, sed, and see genes. The positive isolates were nuc, 100% (17); sea, 35.29% (6); seb, 5.88% (1); sec, 5.88% (1); sed, 29.41% (5); and see, 47.06% (8). The isolates were also tested with 2 enzyme immunoassay diagnostic kits, one for the screening detection of the production of staphylococcal enterotoxins (SEA, SEB, SEC, SED, SEE) and one for the detection of specific enterotoxin produced by each isolate. Seven out of 17 (41.18%) were enterotoxin producers: 7 produced SEA (41.18%), 1 SEB (5.88%), 1 SEC (5.88%), 5 SED (29.41%), and 6 SEE (35.29%). To further characterize the isolates, they were analyzed by the Kirby Bauer test for susceptibility to 13 antimicrobials (ampicillin, ciprofloxacin, kanamycin, tetracycline, gentamicin, methicillin, nalidixic acid, erythromycin, amoxicillin/clavulanic acid, streptomycin, vancomycin, neomycin, and enrofloxacin), and we detected resistance to ampicillin (52.94%), nalidixic acid (70.59%), erythromycin (5.88%), and amoxicillin/clavulanic acid (17.65%). The isolates were sensitive to the main classes of antimicrobials used for the treatment of bovine subclinical mastitis. The presence of enterotoxin-producing isolates of S. aureus in bovine milk means that a temperature abuse or a breakdown in the thermal treatment of the milk could present a food safety risk, particularly if all enterotoxigenic isolates could potentially produce SEA in milk.

Detection and characterization of methicillin-resistant and susceptible coagulase-negative staphylococci in milk from cows with clinical mastitis in Tunisia.

OBJECTIVES: This study investigated prevalence of methicillin-resistant (MR) and methicillin-susceptible (MS) coagulase-negative staphylococci (CNS) and the implicated mechanisms of resistance and virulence in milk of mastitis cows. In addition, the presence of SCCmec type was analyzed in MR Staphylococcus epidermidis (MRSE). RESULTS: Three hundred milk samples from cows with clinical mastitis were obtained from 30 dairy farms in different regions of Tunisia. Sixty-eight of the 300 tested samples contained CNS strains. Various CNS species were identified, with Staphylococcus xylosus being the most frequently found (40%) followed by Staphylococcus warneri (12%). The mecA gene was present in 14 of 20 MR-CNS isolates. All of them were lacking the mecC gene. The SCCmecIVa was identified in four MRSE isolates. Most of CNS isolates showed penicillin resistance (70.6%) and 58.3% of them carried the blaZ gene. MR-CNS isolates (n = 20) showed resistance to erythromycin, tetracycline and trimethoprim-sulfametoxazole harboring different resistance genes such us erm(B), erm(T), erm(C), mph(C) or msr(A), tet(K) and dfr(A). However, a lower percentage of resistance was observed among 48 MS-CNS isolates: erythromycin (8.3%), tetracycline (6.2%), streptomycin (6.2%), clindamycin (6.2%), and trimethoprim-sulfametoxazole (2%). The Inu(B) gene was detected in one Staphylococcus xylosus strain that showed clindamycin resistance. The virulence gene tsst-1 was observed in one MR-CNS strain. DISCUSSION: Coagulase-negative staphylococci containing a diversity of antimicrobial resistance genes are frequently detected in milk of mastitis cows. This fact emphasizes the importance of identifying CNS when an intramammary infection is present because of the potential risk of lateral transfer of resistant genes among staphylococcal species and other pathogenic bacteria.