Clinical Bacteriology Laboratory Research Articles

Results of a 10-month Study on the Species Spectrum and Antimicrobial Resistance of Microorganisms in a Tertiary Care Hospital

Introduction. Antimicrobial resistance (AMR) remains a global issue requiring a comprehensive approach to address it. One of the methods to mitigate AMR is the microbiological monitoring of antibiotic-resistant strains of microorganisms within the framework of infection control with regular monitoring of resistance patterns. The aim of thе study. To determine the species diversity and antimicrobial resistance of microorganisms isolated in a tertiary care hospital over a 10-month period. Materials and methods. The study was conducted at the Clinical Bacteriology Laboratory of the Regional Center for Laboratory Diagnostics, Communal enterprise ″Yuri Semenyuk Rivne Regional Clinical Hospital″ of Rivne Regional Council (Rivne, Ukraine) over 10 months (July 2023 - April 2024). From 8.300 patients, 12.463 biological samples were collected, from which 16.618 strains of microorganisms were isolated and studied. Pure culture isolation was performed using bacteriological methods with nutrient media. Species identification and antimicrobial susceptibility testing were carried out using a semi-automated bacteriological analyzer ID/AST system D2mini with plates produced by Zhuhai DL Biotech Co. Ltd., in accordance with EUCAST 2023-2024 guidelines. Results. During the 10 months of observation, 16.618 strains of microorganisms were identified and their antimicrobial resistance levels were determined. Of these, 64.4 % were gram-negative bacteria, 32.5 % were gram-positive microorganisms, and 3.0 % were yeast fungi. The most common gram-negative bacteria were Klebsiella pneumoniae (40.8 %), Pseudomonas aeruginosa (18.2 %), and Acinetobacter baumannii (14.4 %). Among the gram-positive bacteria, Staphylococcus aureus (39.5 %), Staphylococcus epidermidis (15.5 %), and Staphylococcus haemolyticus (14.1 %) were predominant. The most common yeast fungi of the Candida genus were Candida albicans (60.5 %), Candida guilliermondii (12.5 %), and Candida tropicalis (8.9 %). High resistance was found to second-generation cephalosporins (81.4 % resistant strains to cefalexin) and fourth-generation cephalosporins (67.2 % resistant to cefepime), as well as aminopenicillins (amoxicillin-clavulanate – 76.1 %, ampicillin – 75.1 %). High sensitivity was noted to antibiotics of the tetracycline group (78.5 % to tetracycline), glycylcyclines (93.5 % to tigecycline), oxazolidinones (93.5 % to tigecycline), and lincosamides (80.8 % to clindamycin). Conclusions. A total of 16.618 strains of microorganisms were identified, with 64.4 % being gram-negative bacteria, 32.5 % gram-positive microorganisms, and 3.0 % yeast fungi. High resistance was found to second- and fourth-generation cephalosporins and aminopenicillins. High sensitivity was detected to antibiotics of the tetracycline, glycylcycline, oxazolidinone, and lincosamide groups.

Development and evaluation of an artificial intelligence for bacterial growth monitoring in clinical bacteriology.

In clinical bacteriology laboratories, reading and processing of sterile plates remain a significant part of the routine workload (30%-40% of the plates). Here, an algorithm was developed for bacterial growth detection starting with any type of specimens and using the most common media in bacteriology. The growth prediction performance of the algorithm for automatic processing of sterile plates was evaluated not only at 18-24 h and 48 h but also at earlier timepoints toward the development of an early growth monitoring system. A total of 3,844 plates inoculated with representative clinical specimens were used. The plates were imaged 15 times, and two different microbiologists read the images randomly and independently, creating 99,944 human ground truths. The algorithm was able, at 48 h, to discriminate growth from no growth with a sensitivity of 99.80% (five false-negative [FN] plates out of 3,844) and a specificity of 91.97%. At 24 h, sensitivity and specificity reached 99.08% and 93.37%, respectively. Interestingly, during human truth reading, growth was reported as early as 4 h, while at 6 h, half of the positive plates were already showing some growth. In this context, automated early growth monitoring in case of normally sterile samples is envisioned to provide added value to the microbiologists, enabling them to prioritize reading and to communicate early detection of bacterial growth to the clinicians.

Continual Improvement in Clinical Bacteriology Laboratory with Quality Indicators: A Retrospective Observational Study

Introduction: Healthcare management is undergoing significant changes with the evolution of new and re-emerging infections. A clinical microbiologist plays an important role in giving an accurate and timely report to the clinicians. Quality Indicators (QIs) act as a measure of the quality of services offered by the laboratory and are tools to monitor and evaluate the laboratory’s performance throughout the Total Testing Process (TTP). Aim: To measure the performance of the clinical bacteriology laboratory using QIs. Materials and Methods: A retrospective study was conducted in the Department of Microbiology at Lokmanya Tilak Municipal Medical College and General Hospital, Mumbai, Maharashtra, India. The study evaluated QIs from the records of 94,624 samples received in the bacteriology section of the clinical Microbiology laboratory between January 2018 and March 2021. Data analysis was conducted over a six-month period from December 2021 to May 2022. In 2018, one QI was identified for each phase, with an additional QI added in each phase to the pre-existing QI in 2019. In 2020, a QI was added in the preanalytical phase only. In 2021, the acceptable limit for one preanalytical QI was reduced from 2% to 1%. Data analysis was performed using an Excel sheet. Results: Data from records of 94,624 clinical bacteriology samples collected over 39 months were analyzed retrospectively. The preanalytical indicators included the number of samples rejected (135, 0.14%) and the number of requisition forms with three patient identifiers (59,645, 93.95%). Analytical phase QIs consisted of the average External Quality Assurance Scheme (EQAS) performance score (97.44% from January 2018 to March 2021) and outliers in the Internal Quality Control (IQC) (25 from January 2019 till March 2021). Failures in the IQC were not assessed in 2018. Postanalytical phase QIs included Turnaround Time (TAT) (average of 2.55 days for aerobic growth) and reporting time for critical alerts, which was within 24 hours of alert finding (100% for smear and culture-positive results). Conclusion: Regular monitoring of QIs helps to identify potential errors. This laboratory chose to analyse and monitor its processes using practically feasible QIs. It was found that the laboratory consistently maintained its performance throughout the study period.

Strengthening clinical bacteriology laboratory diagnostics to combat sepsis and antimicrobial resistance in Benin: a train-the-trainer approach.

Improved laboratory diagnostics is needed to support sepsis diagnosis and combat increasing antibiotic resistance in Benin. We trained clinical laboratory experts and technicians to improve their skills in accurate and up-to-date diagnostics. A Train-the-Trainer (TtT) approach was used to design the course that combines theoretical and practical laboratory skills, specifically addressing the knowledge gaps we had previously identified in our national survey. Pedagogical methods were student-centered, including peer learning, use of online materials, practical laboratory work and pre-and post-course tests. We first trained 10 trainers who in turn trained 40 laboratory technicians from across the country, from both public and private clinical and veterinary laboratories. The trainers also prepared standard operation procedures for blood culture and antibiotic susceptibility testing based on international standards. Three months after the training, follow-up visits were made to the laboratories where the implementation of the new skills was evaluated. The progress of the participants observed during the course and the implementation of the new skills afterwards proved the training to be effective. The professional networks created during the training, the empowerment that utilizes local knowledge resources, and the government support for our initiative can be expected to bring sustainability to the initiative and support the participation of Beninese laboratories in international surveillance programs in the future.

An epidemiological study of the predictors of multidrug resistance and methicillin resistance among Staphylococcus spp. isolated from canine specimens submitted to a diagnostic laboratory in Tennessee, USA.

Understanding drivers of multidrug resistance (MDR) and methicillin resistance, which have increased among canine staphylococcal isolates, is essential for guiding antimicrobial use practices. Therefore, the objective of this study was to identify predictors of MDR and methicillin resistance among Staphylococcus spp. commonly isolated from canine clinical specimens. This retrospective study used records of canine specimens submitted to the University of Tennessee College of Veterinary Medicine Clinical Bacteriology Laboratory for bacterial culture and antimicrobial susceptibility testing between 2006 and 2017. Records from 7,805 specimens positive for the following Staphylococcus species were included for analysis: Staphylococcus pseudintermedius, Staphylococcus aureus, Staphylococcus coagulans (formerly Staphylococcus schleiferi subspecies coagulans), and Staphylococcus schleiferi (formerly S. schleiferi subsp. schleiferi). Generalized linear regression models were fit using generalized estimating equations (GEE) to identify predictors of MDR (defined as resistance to three or more antimicrobial classes) and methicillin resistance among these isolates. Multidrug resistance (42.1%) and methicillin resistance (31.8%) were relatively common. Isolates from skeletal (joint and bone) specimens had the highest levels of MDR (51.3%) and methicillin resistance (43.6%), followed by cutaneous specimens (45.8% multidrug-resistant, 37.1% methicillin resistant). Staphylococcus species, specimen site, and clinical setting were significant (p<0.01) predictors of both outcomes. Compared to S. pseudintermedius, S. schleiferi had higher odds of methicillin resistance, while S. coagulans and S. schleiferi had lower odds of MDR. The odds of both MDR and methicillin resistance for isolates from hospital patient specimens were significantly higher than those from referral patients for urine/bladder and otic specimens. Odds of MDR among isolates from skeletal specimens of hospital patients were also higher than those of referral patients. Staphylococcus isolates in this study had substantial levels of MDR and methicillin resistance. Differences in the odds of these outcomes between referral and hospital patient isolates did not persist for all specimen sites, which may reflect differences in diagnostic testing and antimicrobial use practices with respect to body site or system. Judicious antimicrobial use, informed by culture and susceptibility testing, is important to limit treatment failures and curb selection pressure.

Evaluation of InTray Cassettes Directly from Blood Cultures for the Diagnosis of Sepsis in Clinical Bacteriology Laboratories as an Alternative to Classic Culture Media.

Culture media is fundamental in clinical bacteriology for the detection and isolation of bacterial pathogens. However, in-house media preparation could be challenging in low-resource settings. InTray® cassettes (Biomed Diagnostics) could be a valid alternative as they are compact, ready-to-use media preparations. In this study, we evaluate the use of two InTray media as a subculture alternative for the diagnosis of bloodstream infections: the InTray® Müller-Hinton (MH) chocolate and the InTray® Colorex™ Screen. The InTray MH chocolate was evaluated in 2 steps: firstly, using simulated positive blood cultures (reference evaluation study), and secondly, using positive blood cultures from a routine clinical laboratory (clinical evaluation study). The Colorex Screen was tested using simulated poly-microbial blood cultures. The sensitivity and specificity of the InTray MH chocolate were respectively 99.2% and 90% in the reference evaluation study and 97.1% and 88.2% in the clinical evaluation study. The time to detection (TTD) was ≤20 h in most positive blood cultures (99.8% and 97% in the two studies, respectively). The InTray® MH Chocolate agar showed good performance when used directly from clinical blood cultures for single bacterial infections. However, mixed flora is more challenging to interpret on this media than on Colorex™ Screen, even for an experienced microbiologist.

Validation of Three MicroScan® Antimicrobial Susceptibility Testing Plates Designed for Low-Resource Settings.

Easy and robust antimicrobial susceptibility testing (AST) methods are essential in clinical bacteriology laboratories (CBL) in low-resource settings (LRS). We evaluated the Beckman Coulter MicroScan lyophilized broth microdilution panel designed to support Médecins Sans Frontières (MSF) CBL activity in difficult settings, in particular with the Mini-Lab. We evaluated the custom-designed MSF MicroScan Gram-pos microplate (MICPOS1) for Staphylococcus and Enterococcus species, MSF MicroScan Gram-neg microplate (MICNEG1) for Gram-negative bacilli, and MSF MicroScan Fastidious microplate (MICFAST1) for Streptococci and Haemophilus species using 387 isolates from routine CBLs from LRS against the reference methods. Results showed that, for all selected antibiotics on the three panels, the proportion of the category agreement was above 90% and the proportion of major and very major errors was below 3%, as per ISO standards. The use of the Prompt inoculation system was found to increase the MIC and the major error rate for some antibiotics when testing Staphylococci. The readability of the manufacturer’s user manual was considered challenging for low-skilled staff. The inoculations and readings of the panels were estimated as easy to use. In conclusion, the three MSF MicroScan MIC panels performed well against clinical isolates from LRS and provided a convenient, robust, and standardized AST method for use in CBL in LRS.

Patterns of antimicrobial, multidrug and methicillin resistance among Staphylococcus spp. isolated from canine specimens submitted to a diagnostic laboratory in Tennessee, USA: a descriptive study

BackgroundMultidrug- and methicillin-resistant staphylococci are both veterinary and public health concerns due to their zoonotic potential. Therefore, the objective of this study was to investigate patterns of antimicrobial, multidrug, and methicillin resistance among four Staphylococcus spp. commonly isolated from canine clinical specimens submitted to the Clinical Bacteriology Laboratory at the University of Tennessee College of Veterinary Medicine (UTCVM).MethodsResults of antimicrobial susceptibility testing and mecA polymerase chain reaction (PCR) for isolates of four common Staphylococcus spp. isolates were obtained from the Bacteriology Laboratory at the UTCVM between 01/01/2006 and 12/31/2017. Cochran-Armitage trend test was used to assess temporal trends of antimicrobial resistance (AMR), multidrug resistance (MDR), and methicillin resistance. Kappa test of agreement was used to assess agreement between the results of PCR and disk diffusion tests.ResultsMost of the 7805 isolates were S. pseudintermedius (6453 isolates), followed by S. coagulans (860), S. aureus (330), and S. schleiferi (162). Among S. pseudintermedius isolates, 45.5% were MDR, and 30.8% were methicillin-resistant (MRSP). There was a significant temporal increase in MRSP (p = 0.017). Chloramphenicol resistance increased among both MRSP and methicillin-susceptible (MSSP) isolates (p < 0.0001). Among S. aureus isolates, 40.9% were MDR, 37.4% were methicillin-resistant (MRSA), and the proportion of MRSA isolates increased significantly (p = 0.0480) over time. There was an increasing temporal trend in the proportion of MDR isolates among MSSP (p = 0.0022), but a decrease among MRSP (p < 0.0001) and MRSA (p = 0.0298). S. schleiferi had the highest percentage (56.9%) of methicillin-resistant isolates. Oxacillin disk diffusion was superior to cefoxitin for the detection of mecA-mediated resistance and had almost perfect agreement with mecA PCR assay for S. pseudintermedius (95.4% agreement, kappa (κ) = 0.904; p < 0.0001), S. coagulans (95.6%, κ = 0.913; p < 0.0001) and S. schleiferi (97.7%, κ = 0.945; p < 0.0001). However, cefoxitin disk diffusion was superior to oxacillin disk diffusion and had almost perfect agreement with mecA PCR assay for S. aureus (95.3%, κ = 0.834; p < 0.0001).ConclusionsThe levels of resistance and increasing temporal trends are concerning. These findings have implications for treatment decisions and public health due to the zoonotic potential of staphylococci. Continued surveillance and use of antibiograms to guide clinical decisions will be critical.

Blood culture surveillance in a secondary care hospital in Benin: epidemiology of bloodstream infection pathogens and antimicrobial resistance

BackgroundAlthough global surveillance of antimicrobial resistance (AMR) is considered key in the containment of AMR, data from low- and middle-income countries, especially from sub-Saharan Africa, are scarce. This study describes epidemiology of bloodstream infections and antimicrobial resistance rates in a secondary care hospital in Benin.MethodsBlood cultures were sampled, according to predefined indications, in BacT/ALERT FA Plus and PF Plus (bioMérieux, Marcy-l’Etoile, France) blood culture bottles (BCB) in a district hospital (Boko hospital) and to a lesser extent in the University hospital of Parakou. These BCB were incubated for 7 days in a standard incubator and twice daily inspected for visual signs of growth. Isolates retrieved from the BCB were processed locally and later shipped to Belgium for reference identification [matrix-assisted laser desorption/ionization time-of-flight spectrometry (MALDI-TOF)] and antibiotic susceptibility testing (disk diffusion and E-tests).ResultsFrom October 2017 to February 2020, 3353 BCB were sampled, corresponding to 3140 blood cultures (212 cultures consisting of > 1 BCB) and 3082 suspected bloodstream infection (BSI) episodes. Most of these cultures (n = 2471; 78.7%) were sampled in children < 15 years of age. Pathogens were recovered from 383 (12.4%) cultures, corresponding to 381 confirmed BSI. 340 of these pathogens were available and confirmed by reference identification. The most common pathogens were Klebsiella pneumoniae (n = 53; 15.6%), Salmonella Typhi (n = 52; 15.3%) and Staphylococcus aureus (n = 46; 13.5%). AMR rates were high among Enterobacterales, with resistance to third-generation cephalosporins in 77.6% of K. pneumoniae isolates (n = 58), 12.8% of Escherichia coli isolates (n = 49) and 70.5% of Enterobacter cloacae isolates (n = 44). Carbapenemase production was detected in 2 Escherichia coli and 2 Enterobacter cloacae isolates, all of which were of the New Delhi metallo-beta lactamase type. Methicillin resistance was present in 22.4% of S. aureus isolates (n = 49).ConclusionBlood cultures were successfully implemented in a district hospital in Benin, especially among the pediatric patient population. Unexpectedly high rates of AMR among Gram-negative bacteria against commonly used antibiotics were found, demonstrating the clinical and scientific importance of clinical bacteriology laboratories at this level of care.

Analysis of trends of post-analytical quality indicators in Clinical Bacteriology laboratory: A pilot study from a tertiary care teaching hospital in Uttarakhand, India

ContextImplementation of quality management system (QMS) which encompasses various quality indicators (QIs), can serve as a stepping stone for continuous improvement & help in achieving globally accepted quality standards in a diagnostic laboratory. AimsTo generate preliminary data on trends of post-analytical QIs in Bacteriology section. Settings and designA pilot study was conducted at a tertiary care teaching hospital located in Rishikesh, Uttarakhand. Methods and materialData of the following four quality indicators pertaining to aerobic culture and sensitivity testing of various clinical samples received in Clinical Bacteriology laboratory was compiled and retrospectively analysed: (i) Rate of reporting errors; (ii) Rate of re-dos; (iii) Percentage of reports correlating with clinical diagnosis; (iv) Percentage of adherence to safety precautions by employees working in diagnostics. Statistical analysis usedDescriptive statistics like mean and frequency distribution plots. ResultsThe mean reporting error rate was 0.12 per 1000 tests. It was consistently low from July 2018 to May 2019, after which an overall increasing trend was observed. The mean rate of re-dos was 2.79 per 1000 tests. An overall decreasing trend was observed with maximum rates during the months of December 2017 and January 2018. On an average only 7.86% of the reports co-related with clinical diagnosis. Almost 100% adherence to safety precautions was observed with the exception of two instances of needle stick injuries (NSIs). ConclusionsCommitment of laboratory personnel in adopting, maintaining and analysing QMS data will lead to further strengthening of our existing healthcare system.

Improving Confirmatory Testing for the Antimicrobial Resistance Surveillance Network in Ethiopia

Background: In July 2017, the Ethiopian Public Health Institute (EPHI) launched an antimicrobial resistance (AMR) surveillance network at 4 sentinel laboratories. The National Clinical Bacteriology and Mycology Laboratory (NRL) at EPHI performs monthly confirmatory testing on a subset of isolates submitted by these sites. We assessed the existing confirmatory testing program to identify gaps and develop solutions, including a monitoring and evaluation (M&amp;E) system. Methods: We assembled a technical working group (TWG) of key stakeholders. Laboratory site visits included workflow observation, process mapping, document review, and technologist interviews. Proposed solutions to observed gaps were drafted in formats consistent with their intended application. Feedback from the TWG was incorporated into final drafts. Available AMR network staff members were trained remotely, and they will train remaining staff. Results: Table 1 describes major gaps and solutions identified. Conclusions: Confirmatory testing provides a mechanism to evaluate laboratory testing proficiency, target improvements, and estimate surveillance data quality, yet standardized methods were lacking. Our efforts highlight key components of confirmatory testing programs and provide a model for use in laboratories with similar needs.Funding: NoneDisclosures: None

Improving Report Generation and Delivery System of Microbiological Investigations at MRI – Sri Lanka with Concern to Turn-Around-Time, An Intervention Study

&lt;p&gt;Background: Clinical Bacteriology Laboratory (CBL) of the Medical Research Institute (MRI) is a National Reference Laboratory in Sri Lanka. The service improvement research project aimed at improving the report generation and delivery system. &lt;br /&gt;Methods: The mixed-method applied to study the process. Gap identification and designing of the intervention used a qualitative approach. Quantitative methods measured the effectiveness of the improvements. A package of interventions based on total quality management (TQM) principles designed with all stakeholder participation. &lt;br /&gt;Results: Delay in sample transport resulted from poor communication, lack of established mechanism and weak coordination between the transport unit and wards. The paper-based report generation process was disorganized, unreliable and error-prone. Additionally, the report delivery to Hospitals revolved on the incoming ambulance or government postal service a tied-up, poorly coordinated system. The interventions promoted coordinated communication. WHONET, a free windows based Laboratory Information System and database software, developed by the World Health Organization for microbiology laboratories installed to generate reliable and accurate automated reports. Reports in PDF format delivered to end-users by secured email. Timeliness, accuracy, quality and uniformity of reports improved. Improvement in post-intervention turn-around-time was statistically significant (p&amp;lt;0.05). &lt;br /&gt;Conclusion: The interventions based on TQM principles improved the process of report generation and delivery. Laboratory automation is a feasible and effective solution in the local setting. The end-user satisfaction perceived to be high on the improvements.&lt;/p&gt;

Implementation of the WASPLab™ and first year achievements within a university hospital.

In essence, automation can be driven by several of the following incentives: increased processing capacity of the laboratory, better costs control through processes standardization, optimized traceability, or improved workflows to reduce turnaround times (TAT). This project aims at presenting an overview of the project management and change management with a focus on the major challenges addressed by lab staff and laboratory leadership during the different phases of the implementation of the WASPLab™ in a routine clinical bacteriology laboratory. This paper reports our experience and reviews changes in the bacteriology laboratory at Geneva University Hospitals when shifting to the WASPLab™. Practically, the whole automation process was segmented into different packages (specimen type-based segmentation) allowing sequential validation, staff training, and routine implementation. Such process allowed reaching 90% of the identified "automatable" samples within 1year, including personal training, documentation for accreditation supported by publications, without interrupting routine operations. In addition, we implemented a validated automated solution for antimicrobial disk diffusion susceptibility testing. Structured supervision and accurate monitoring of all the activities related to the automation project including key partners such as IT support, technical committee, and after-sales service guaranteed a swift and timely achievement of the project allowing the improvement of the workflow in routine bacteriology within 1year.

Comparison of the Copan WASPLab incorporating the BioRad expert system against the SIRscan 2000 automatic for routine antimicrobial disc diffusion susceptibility testing

ObjectivesThis study investigated the agreement at the categorical level between the Copan WASPLab incorporating the BioRad expert system against the SIRscan 2000 automatic for antimicrobial disc diffusion susceptibility testing. MethodsThe 338 clinical strains (67 Pseudomonas aeruginosa, 19 methicillin-resistant Staphylococcus aureus, 75 methicillin-sensitive S. aureus and 177 Enterobacterales isolates) analysed in this study were non-duplicate isolates obtained from consecutive clinical samples referred to the clinical bacteriology laboratory at Geneva University Hospitals between June and August 2019. For the WASPLab the inoculum suspension was prepared in strict accordance with the manufacturer's instruction (Copan WASP srl, Brescia, Italy) by adding 2 mL of the 0.5 McFarland primary suspension used for the SIRscan analysis into a sterile tube filled with 4 mL of sterile saline (1:3 dilution). The inoculum (2 × 30 μL loop/spreader) was spread over the entire surface of Mueller–Hinton agar plates according to the AST streaking pattern defined by Copan. The antibiotic discs were dispensed by the WASP and inoculated media were loaded on conveyors for transfer to the automatic incubators. The plates were incubated for 16 h, and several digital images were acquired. Inhibition zone diameters were automatically read by the WASPLab and were adjusted manually whenever necessary. For the SIRscan 2000 automatic, the antimicrobial disc diffusion susceptibility testing was performed according to the EUCAST guidelines. The gradient strip method was used to resolve discrepancies. ResultsThe overall categorical agreement between the compared methods reached 99.1% (797/804; 95% CI 98.2%–99.6%), 99.5% (1029/1034; 95% CI 98.9%–99.8%), and 98.8% (2798/2832; 95% CI 98.3%–99.1%) for P. aeruginosa, S. aureus and the Enterobacterales, respectively. ConclusionsWASPLab incorporating the BioRad expert system provides a fully automated solution for antimicrobial disc diffusion susceptibility testing with equal or better accuracy than other available phenotypic methods.

Diagnostic Bacteriology in District Hospitals in Sub-Saharan Africa: At the Forefront of the Containment of Antimicrobial Resistance.

This review provides an update on the factors fuelling antimicrobial resistance and shows the impact of these factors in low-resource settings. We detail the challenges and barriers to integrating clinical bacteriology in hospitals in low-resource settings, as well as the opportunities provided by the recent capacity building efforts of national laboratory networks focused on vertical single-disease programmes. The programmes for HIV, tuberculosis and malaria have considerably improved laboratory medicine in Sub-Saharan Africa, paving the way for clinical bacteriology. Furthermore, special attention is paid to topics that are less familiar to the general medical community, such as the crucial role of regulatory frameworks for diagnostics and the educational profile required for a productive laboratory workforce in low-resource settings. Traditionally, clinical bacteriology laboratories have been a part of higher levels of care, and, as a result, they were poorly linked to clinical practices and thus underused. By establishing and consolidating clinical bacteriology laboratories at the hospital referral level in low-resource settings, routine patient care data can be collected for surveillance, antibiotic stewardship and infection prevention and control. Together, these activities form a synergistic tripartite effort at the frontline of the emergence and spread of multi-drug resistant bacteria. If challenges related to staff, funding, scale, and the specific nature of clinical bacteriology are prioritized, a major leap forward in the containment of antimicrobial resistance can be achieved. The mobilization of resources coordinated by national laboratory plans and interventions tailored by a good understanding of the hospital microcosm will be crucial to success, and further contributions will be made by market interventions and business models for diagnostic laboratories. The future clinical bacteriology laboratory in a low-resource setting will not be an “entry-level version” of its counterparts in high-resource settings, but a purpose-built, well-conceived, cost-effective and efficient diagnostic facility at the forefront of antimicrobial resistance containment.

English

Antimicrobials have been playing an important role in preventing illness and death associated with infections due to bacteria. However, the emergence and spread of resistance by pathogens have decreased the effectiveness of the commonly prescribed antimicrobials. Intestinal Escherichia coli are among bacterial pathogens that are endowed with such resistance traits because they are important source and reservoir of genes that encode antimicrobial resistance. To determine the antimicrobial resistance profile of fecal isolates of E. coli from diarrheic patients. Stool samples were collected consecutively from 100 individuals who visited Selam Health Center during the study period, April to June 2018. Samples were collected and transported under sterile condition to the National Clinical Bacteriology and Mycology reference Laboratory, Ethiopian Public Health Institute. The samples were streaked on MacConkey agar and incubated overnight at 37°C. E. coli isolates were further confirmed using conventional biochemical tests. Antimicrobial susceptibility status was determined using the disk diffusion method on Mueller Hinton agar as recommended by the Clinical Laboratory Standard Institute. The raw data was compiled and entered to spreadsheet and analysis was done using SPSS Version 20 with p-value ≤0.05 considered statistically significant. Out of the 100 patients, 43 were female and the rest were male. Confirmed E. coli were isolated from 73 individuals. Antimicrobial susceptibility testing showed that E. coli isolated in this study were highly resistant to trimethoprim-sulfamethoxazole 49 (67.1%) and amoxicillin-clavulanic acid 47(64.4 %). No isolates showed resistance to gentamicin and tobramicin. Of all the isolates, 11(15.1%) were multidrug resistant. No association was observed between antimicrobial resistance status and sex of individuals included in this study. However, there was an association between age and resistance patterns. Resistance to commonly prescribed antibiotics among E. coli isolated in this study was high and a considerable proportions of the strains were multidrug resistant. This is an indication for an alarming rate of resistance of intestinal E. coli to first line antimicrobials. To reduce the problem, regular monitoring and education for the community are very important. Key words: Escherichia coli, antibiotic susceptibility, multidrug resistant, Ethiopia, biochemical tests, disk diffusion.

Development of an all-in-one transportable clinical bacteriology laboratory: feedback from testing a Mini-Lab prototype

Development of an all-in-one transportable clinical bacteriology laboratory: feedback from testing a Mini-Lab prototype

Latin American consensus to define, categorize, and report multidrug-resistant, extensively drug-resistant, or pandrug-resistant pathogensConsenso latino-americano para definição, categorização e notificação de patógenos multirresistentes, com resistência ampliada ou panresistentes

RESUMENSe presenta un consenso latinoamericano que permite estandarizar las definiciones de los diferentes niveles de resistencia a los antimicrobianos en bacterias de importancia en salud pública. Se describen los criterios de inclusión y exclusión para las metodologías a utilizar y para los antibióticos a incluir (por disponibilidad, relevancia y existencia de puntos de corte). Como propuesta piloto se eligieron tres microorganismos gramnegativos de gran impacto en el ambiente hospitalario (Klebsiella pneumoniae, Pseudomonas aeruginosa y Acinetobacter spp.). La falta de puntos de corte para ciertos antibióticos (por ejemplo, tigeciclina, fosfomicina y colistina), claves para el tratamiento de infecciones causadas por estos patógenos que presentan multirresistencia o resistencia extendida, llevó a la necesidad de discutir y consensuar puntos de corte provisorios para la vigilancia de la resistencia a estos fármacos. Se abordó y consensuó también el uso de pruebas de sensibilidad alternativas a los métodos aprobados por las guías internacionales, de aplicación más sencilla como pruebas de rutina en los laboratorios de bacteriología clínica. El principal beneficio de este documento es proporcionar a los laboratorios latinoamericanos un marco estandarizado y consensuado para la identificación y la vigilancia constante y unificada de microorganismos resistentes. Las recomendaciones incluidas en este documento son el resultado consensuado por los representantes de los laboratorios nacionales de referencia de los países que integran la Red Latinoamericana de Vigilancia de la Resistencia a los Antibióticos coordinada por la Organización Panamericana de la Salud.

Isolation and partial characterization of Streptococcus suis from clinical cases in cattle.

Sixteen isolates of gram-positive, coccoid bacteria were obtained from clinical cases of diverse conditions in cattle and identified as Streptococcus suis using 16S ribosomal DNA gene sequencing and other bacterial identification methods. None of the isolates could be assigned to any of the known S. suis capsular types. Virulence-associated gene profiling that targeted muramidase-released protein, extracellular protein factor, suilysin, 89-kb pathogenicity island, and arginine deiminase ( arcA) genes were negative except for 1 isolate that was arcA positive. The arcA-positive isolate caused severe widespread lesions, including multiorgan suppurative and hemorrhagic inflammation in the meninges, lung, liver, spleen, lymph nodes, and serosae of heart and intestines. The other isolates were primarily associated with meningitis, bronchopneumonia, and multifocal acute necrotizing hepatitis. The isolates differed from each other by 4-6 fragments when examined by pulsed-field gel electrophoresis, indicating they are possibly related. The isolates were susceptible to ampicillin, penicillin, and tiamulin. Resistance was noted to sulfadimethoxine (93%), oxytetracycline (86%), chlortetracycline (86%), neomycin (67%), tilmicosin (47%), clindamycin (47%), enrofloxacin (33%), gentamicin (13%), florfenicol (7%), trimethoprim-sulfamethoxazole (7%), and spectinomycin (53%). Multi-drug resistance (defined as resistance to at least 1 agent in 3 or more antimicrobial classes) was detected in 67% of the isolates. The pathology observations provide evidence that S. suis may be an important pathogen of bovine calves. S. suis is an agent that clinical bacteriology laboratories should consider when dealing with cases involving cattle.

Real-Time Fluorescence Loop Mediated Isothermal Amplification for the Detection of Acinetobacter baumannii

BackgroundDetection of Acinetobacter baumannii has been relying primarily on bacterial culture that often fails to return useful results in time. Although DNA-based assays are more sensitive than bacterial culture in detecting the pathogen, the molecular results are often inconsistent and challenged by doubts on false positives, such as those due to system- and environment-derived contaminations. In addition, these molecular tools require expensive laboratory instruments. Therefore, establishing molecular tools for field use require simpler molecular platforms. The loop-mediated isothermal amplification method is relatively simple and can be improved for better use in a routine clinical bacteriology laboratory. A simple and portable device capable of performing both the amplification and detection (by fluorescence) of LAMP in the same platform has been developed in recent years. This method is referred to as real-time loop-mediated isothermal amplification. In this study, we attempted to utilize this method for rapid detection of A. baumannii.Methodology and Significant FindingsSpecies-specific primers were designed to test the utility of this method. Clinical samples of A. baumannii were used to determine the sensitivity and specificity of this system compared to bacterial culture and a polymerase chain reaction method. All positive samples isolated from sputum were confirmed to be the species of Acinetobacter by 16S rRNA gene sequencing. The RealAmp method was found to be simpler and allowed real-time detection of DNA amplification, and could distinguish A. baumannii from Acinetobacter calcoaceticus and Acinetobacter genomic species 3. DNA was extracted by simple boiling method. Compared to bacterial culture, the sensitivity and specificity of RealAmp in detecting A. baumannii was 98.9% and 75.0%, respectively.ConclusionThe RealAmp assay only requires a single unit, and the assay positivity can be verified by visual inspection. Therefore, this assay has great potential of field use as a molecular tool for detection of A. baumannii.