Antimicrobial susceptibility testing for Helicobacter pylori in times of increasing antibiotic resistance

The gram-negative bacterium Helicobacter pylori(H.pylori)causes chronic gastritis,gastric and duodenal ulcers,gastric cancer and mucosa-associated lymphoid tissue lymphoma.Treatment is recommended in all symptomatic patients.The current treatment options for H.pylori infection are outlined in this review in light of the recent challenges in eradication success,largely due to the rapid emergence of antibiotic resistant strains of H.pylori.Antibiotic resistance is a constantly evolving process and numerous studies have shown that the prevalence of H.pylori antibiotic resistance varies significantly from country to country,and even between regions within the same country.In addition,recent data has shown that previous antibiotic use is associated with harbouring antibiotic resistant H.pylori.Local surveillance of antibiotic resistance is warranted to guide clinicians in their choice of therapy.Antimicrobial resistance is assessed by H.pylori culture and antimicrobial susceptibility testing.Recently developed molecular tests offer an attractive alternative to culture and allow for the rapid molecular genetic identification of H.pylori and resistance-associated mutations directly from biopsy samples or bacterial culture material.Accumulating evidence indicates that surveillance of antimicrobial resistance by susceptibility testing is feasible and necessary to inform clinicians in their choice of therapy for management of H.pylori infection.

Importance of antimicrobial susceptibility testing for the management of eradication in Helicobacter pylori infection

The management of Helicobacter pylori(H. pylori) infection treatment differs from the common treatment protocol for other infectious diseases. Because culture-or molecular-guided approaches face several practical issues, such as the invasive procedures required to obtain gastric biopsy specimens and the lack of availability of routine laboratory testing in some places, H. pylori treatment includes the administration of two or three empirically selected antibiotics combined with a proton pump inhibitor rather than evidence-based eradication treatment. The efficacy of empirical therapy is decreasing, mostly due to increasing multiple resistance. Multiresistance to levofloxacin, clarithromycin, and metronidazole, which are commonly used in empirical treatments, appears to have increased in many countries. Mutations play a primary role in the antimicrobial resistance of H. pylori, but many different mechanisms can be involved in the development of antibiotic resistance. Determining and understanding these possible mechanisms might allow the development of new methods for the detection of H. pylori and the determination of antimicrobial resistance. A treatment based on the detection of antimicrobial resistance is usually more effective than empirical treatment. Nevertheless, such an approach before treatment is still not recommended in the Maastricht guidelines due to the difficulty associated with the routine application of available cultureor molecular-based susceptibility tests, which are usually administered in cases of treatment failure. The management of first and rescue treatments requires further research due to the steadily increase in antimicrobial resistance.

Expert Consensus on Polymyxin Antimicrobial Susceptibility Testing and Clinical Interpretation

The polymyxins are important antimicrobial agents against antibiotic-resistant gram-negative bacilli.In 2020,the Clinical and Laboratory Standards Institute modified the clinical breakpoints for polymyxin susceptibility test by eliminating the"susceptible"interpretive category,only reporting intermediate(≤2 mg/L)and resistant(≥4 mg/L).However,the European Committee on Antimicrobial Susceptibility Testing recommended the use of clinical breakpoints of W2 mg/L as susceptible and>2 mg/L as resistant.The first-line laboratorians and clinicians in China have been perplexed by the inconsistence of international polymyxin clinical breakpoints and discouraged by the difficulty of conducting polymyxin susceptibility testing.Therefore,it is urgently needed to make it clear for the laboratorians in China to know how to accurately carry out polymyxin susceptibility testing and standardize the interpretation of susceptibility testing results.To this end,the experts from relevant fields were convened to formulate this consensus statement on the testing and clinical interpretation of polymyxin susceptibility.Relevant recommendations are proposed accordingly for laboratorians and clinicians to streamline their daily work.

Threefold Increase in the Number of Drug Resistant TB Cases after Introduction of Universal Drug Susceptibility Testing: Experiences from Two South India Districts

Background: In India, tuberculosis (TB) is a major public health problem, and the advent of drug resistance TB (DR-TB) has worsened the situation. The Revised National TB Control Programme (RNTCP) has introduced universal drug susceptibility testing (UDST) for all diagnosed TB cases in 2018. We conducted this study to know the advantage of implementing UDST when compared to selective testing existent in 2017 on key diagnostic cascade parameters and to identify the challenges in the implementation of UDST. Methods: The study was conducted in two districts of Karnataka, India during January 2017-December 2018. The quantitative part consisted of before-and-after design and the qualitative part consisted of descriptive design. Results: In 2017 (during selective testing/“before” period) out of the 2440 TB patients, 80 (3%) were diagnosed with Isoniazid and Rifampicin resistance patients;in contrast in 2018 (during UDST/“after” period) of the 5129 TB patients 258 (5%) were diagnosed with Isoniazid and Rifampicin resistance. However, the proportion of eligible patients tested for rifampicin resistance during the “after” period was 60% when compared to 100% during the “before” period and median turnaround time for testing was also longer during the “after” period when compared to the “before” period (32.5 days vs 27.5 days). Major reasons for these two gaps were found to be difficulties in collecting sputum specimens and transportation. Conclusion: The rollout of UDST has led to a three-fold increase in a number of DR-TB cases detected in the region. There is a need for the programme to increase the proportion tested for DST by increasing the laboratory capacity and address the challenges in sputum collection and transportation.

Assessment of Clinical Presentation, Performance of Diagnostic Methods and Antibiotic Susceptibility Testing for Salmonella among Patients Attending Kangema Sub-County Hospital, Kenya

Background: Typhoid disease remains a major public health problem globally, especially in developing countries in sub-Saharan Africa. Symptoms associated with typhoid disease mimic those of other febrile illnesses and are thus difficult to make an accurate diagnosis. A confirmed diagnosis requires the determination or isolation of the bacteria in well-equipped laboratories. Developing countries are faced with a huge limitation of the laboratory infrastructure to diagnose typhoid disease, which would otherwise guide in treating, managing, controlling, and halting the spread of drug resistant mutants. Objective: This study, therefore, was aimed at determining the clinical presentation, performance of diagnostic tests and antibiotic susceptibility testing of Salmonella among adults attending Kangema Sub-County Hospital. Study Population: The study population was residents of Kangema Sub-County in Murang’a County, Kenya while the target population was adults. Methods: The study adopted a cross-sectional study design that employed a systematic random sampling procedure. The study took place between April and June 2021. The sample size was 97 respondents who all consented and were enrolled in the study. Interviewing the respondents was carried out by administering structured questionnaires to collect quantitative data. Stool samples were obtained and cultured in Cary Blair transport media and then cultured in appropriate media at the Murang’a County Referral Hospital Laboratory. A rapid Salmonella Antigen (SAT) test was also performed on all the stool samples. Data Analyses: Word Statistics and Data (STATA) v 13 was used for statistical analysis. Results: The prevalence of Typhoid Fever was at 6.2% (95% CI) which included S. Typhi (n = 1;16.7%) and S. Paratyphi B (n = 5;83.3%). No isolate showed resistance to Ciprofloxacin. The sensitivity of SAT is 100% and a specificity of 98.9% with a kappa statistic of almost perfect agreement (0.9641) with culture. Patients who had fever p = 0.001, abdominal distention p = 0.028, diarrhoea p = 0.038, loose or watery stool p = 0.021 and mild general condition p = 0.02 remained independently associated with Salmonella infection. Conclusion: Typhoid Fever being endemic, laboratory diagnosis was a key for confirmation after clinical diagnosis. SAT can accurately be used to detect the disease where culture is unavailable. However, antibiotic sensitivity tests were crucial when determining the drug of choice as Salmonella isolates were multi-drug resistant. Establishment of prescribing antimicrobial policies and guidelines can periodically monitor the antibiogram patterns.

Identification and Susceptibility Test of Ralstonia solanacearum Isolated from White Burley

[ Objective ] The paper was to isolate and identify Ralstonia solanacearum from white burley, and determine its susceptibility to 6 fungicides. [ Mcth- od] Using the combination method of semiselective medium （PCCG） and apolymerase chain reaction （PCR） technique, R. solanacearum in stalk of white burley from Dazhou City in Sichnan Province was isolated, and its biochemical type was identified. Through susceptibility test, the susceptibility of R. solanacearum to bismerthiazol, ethylicin, streptomycin, lime sulfur, 47% polylysine, 99% kojic acid was studied in laboratory. [Result] A total of 23 strains OfR. solanacearum were isolated, all belonging to biochemical type Ill. R. solanacearum obtained in the test were more susceptible to ethylicin, streptomycin and bismerthiazol, and ethylicin had good control effect against R. solanacearum with ECso of 0.086 ml/L. [ Conclusion ] The study provide theoretical basis for control of R. solanaceanon in white burley.

Oblique Y-groove cracking test of the welding cold cracking susceptibility of domestic X-70 pipeline steel

Gas Transmitting From West to East Project' is significant. It should ensure the welding quality and safety of pipeline. The task is very arduous to guarantee the quality of the project in the condition of long line, complex weather and geology features. In this paper, the welding cold cracking susceptibility of domestic X 70 pipeline steel adopted by the project, which is one of the most interesting questions of welding quality about petrol pipeline, was studied by means of oblique Y groove cracking test. The crack ratio of surface and section was tested under the conditions of different welding materials and preheat temperature .The thickness of plate steel was 14.7 mm and 10.3 mm . The results reveal that X 70 pipeline steel has good crack resistance. The research has important value for the construction of large scale pipeline engineering and the application of domestic X 70 pipeline steel.

Detection of Beta-Lactamase and Extended-Spectrum Beta-Lactamase of Pathogens Isolated from Pig and Chicken and Their Antibiotic Susceptibility Test

The antibacterial activity of beta-lactam antibiotics or their combinations with inhibitor sulbactum against non-lactamase- producing strains, lactamase-producing and ESBLs-producing isolates was evaluated with twofold dilution method after pathogens isolated from pigs and chickens were detected, respectively, for beta-lactamase and extended-spectrum beta- lactamases （ESBLs）, The results revealed that most of 43 clinically isolated strains could produce beta-lactamase and 3 strains of shigella isolated from chicken samples produced ESBLs. All of 30 lactamase-producing strains isolated and only one of 16 non-lactamase-producing strains were resistant to amoxicillin and ampicillin. MICs of ampicillin against lactamaseproducing isolates decreased 10-40 and 10-20 times respectively, when it was conbined with sulbactam at ration of 1：2 and 1：4. All clinical isolates were susceptible to third-generation cephalosporins. The MICs of third-generation cephalosporins against lactamase-producing isolates did not change when they were conbined with sulbactam. MICs of ceftiofur and ceftriaxone against ESBLs-producing isolates decreased 2-4 times when they were conbined with sulbactam.

Drug Susceptibility Test of Volatile Oil of Artemisiaargyi to Avian Pathogenic Escherichia coli

The volatile oil of Artemisia argyi was extracted by ultrasonic assisted extraction, and the extraction rate of volatile oil was 0.68%. Thevolatile oil of A. argyi was emulsified with 1% Tween-80, and drug susceptibility test was conducted with avian Escherichia coli. The results showedthat the volatile oil of A. argyi had antibacterial effect against avian E. coli, and the minimal inhibitory concentration was 50 mg/mL. Taking sixcommon antibiotics as the control, drug susceptibility test was conducted with volatile oil of A. argyi. The results showed that 10 strains of E. coliwere sensitive to the volatile oil of A. argyi, three of which had different degrees of resistance and one had the tendency of resistance.

Isolation,Identification and Antibiotics Susceptibility Test of Citrobacter freundii from Procambarus clarkia

This experiment was conducted to clarify species and drug resistance of pathogen from the diseased Procambarus clarkia. Pathogenic bacteria from hepatopancreas of the diseased P. clarkia were examined using conventional methods,and then were isolated. The further tests and analysis of the isolated strain were developed,including the regression experiment to P. clarkia,the morphology,physiological and biochemical characteristics,sequence analysis of their 16 S rRNA and gyr B genes,and the susceptibility test to antibiotics. Large colonies with similar morphology and color were obtained. Strain X120523 was identified as Citrobacter freundii,proved to have strong pathogenicity,and was susceptible to quinolones and aminoglycosides.

A comparative study on antimicrobial susceptibility of uroculturome of humans in health and urinary tract infections

Background:The uroculturome indicates the profile of culturable microbes inhabiting the urinary tract,and it is often required to do a urine culture to find an effective antimicrobial to treat urinary tract infections(UTIs).Methods:This study targeted to understand the profile of culturable pathogens in the urine of apparently healthy(128)and humans with clinical UTIs(161)and their antimicrobial susceptibility.All the urine samples were analyzed to quantify microbial load and determine the diversity and antimicrobial susceptibility of microbes following standard microbiological methods.Results:In urine samples from UTI cases,microbial counts were 1.2×10^(4)±6.02×10^(3) colony-forming units(cfu)/mL,while in urine samples from apparently healthy humans,the average count was 3.33±1.34×10^(3) cfu/mL.In eight samples(six from UTI cases and two from apparently healthy people,Candida(C.albicans 3,C.catenulata 1,C.krusei 1,C.tropicalis 1,C.parapsiplosis 1,C.gulliermondii 1)and Rhizopus species(1)were detected.Candida krusei was detected only in a single urine sample from a healthy person and C.albicans was detected both in urine of healthy and clinical UTI cases.Gram-positive(G+ve)bacteria were more commonly(Odds ratio,1.98;CI99,1.01-3.87)detected in urine samples of apparently healthy humans,and Gram-negative(G−ve)bacteria(Odds ratio,2.74;CI99,1.44-5.23)in urines of UTI cases.From urine samples of 161 UTI cases,a total of 90 different types of microbes were detected and,73 samples had only a single type of bacteria.In contrast,49,29,3,4,1,and 2 samples had 2,3,4,5,6 and 7 types of bacteria,respectively.The most common bacteria detected in urine of UTI cases was Escherichia coli(52 samples),in 20 cases as the single type of bacteria,other 34 types of bacteria were detected in pure form in 53 cases.From 128 urine samples of apparently healthy people,88 types of microbes were detected either singly or in association with others,from 64 urine samples only a single type of bacteria was detected while 34,13,3,11,2 and 1 sample yielded 2,3,4,5,6 and seven types of microbes,respectively.In the urine of apparently healthy humans too,E.coli was the most common bacteria,(10 samples)followed by Staphylococcus haemolyticus(9),S.intermedius(5),and S.aureus(5),and similar types of bacteria also dominated in cases of mixed occurrence,E.coli was detected in 26,S.aureus in 22 and S.haemolyticus in 19 urine samples,respectively.G+ve bacteria isolated from urine samples’irrespective of health status were more often(P<0.01)resistant than G−ve bacteria to ajowan oil,holy basil oil,cinnamaldehyde,and cinnamon oil,but more susceptible to sandalwood oil(P<0.01).However,for antibiotics,G+ve were more often susceptible than G−ve bacteria to cephalosporins,doxycycline,and nitrofurantoin.Conclusion:The study concludes that to understand the role of good and bad bacteria in the urinary tract microbiome more targeted studies are needed to discern the isolates at the pathotype level.Further,the study suggests the use of antibiotics by observing good antibiotic stewardship following antibiotic susceptibility testing only.

Isolation and Identification of Four Strains of Salmonella and Drug Susceptibility Test

In order to isolate and identify the pathogenic bacteria causing dead chickens in a chicken farm in Qinhuangdao area, the liver, heart and other organs of dead chickens suspected of salmonella disease were collected aseptically, and streaked on SS agar medium and chromagar medium. Transparent colonies were observed on SS agar medium, and purple and transparent colonies on CAS medium. The isolate was conducted purification, staining microscopy, biochemical tests, and 16 S rRNA sequence analysis, and the results showed that four strains of the isolated bacte-ria were salmonella. The 16 S rRNA sequence analysis of four strains of salmonella showed that the isolates shared more than 99% homology. Drug susceptibility test was performed using paper method, and the results showed that most of the strains were resistant to tilmicosin, cefradine and sul-famethoxazole, but were sensitive to ceftriaxone.

Isolation, Identification and Drug Susceptibility Test of a Strain of Escherichia coli Causing Fox Pneumonia

[Objective] The paper was to determine the pathogen causing fox pneumonia in a breeding factory in Changli County.[Method]Through autopsy, a dominant strain was isolated from the lung of dead foxes, which was then performed Gram staining, 16 S rRNA sequence analysis and biochemical identification.[Result] The strain was negative in Gram staining, and was identified as E. coli through 16 S rRNA sequence analysis and biochemical identification. Drug susceptibility test was conducted using 15 kinds of drug susceptibility papers. The E. coli was sensitive to florfenicol, enrofloxacin, ceftriaxone, norfloxacin;intermediately sensitive to amikacin, gentamicin;and strongly resistant to penicillin, ampicillin,cefradine, sulfamethoxazole, lincomycin, streptomycin and amoxicillin.[Conclusion] It is difficult to treat E. coli causing fox pneumonia with traditional antibiotics clinically.

Susceptibility to Fluconazole and Ketoconazole of Candida spp. Isolated from Primary and Episodic Vulvovaginites by E-Test (São Paulo, SP, Brazil)

Purpose: To evaluate the profile of in vitro susceptibility of yeasts isolated from cases of primary and espisodic vulvovaginitis to two antifungal agents. Methods: 40 Candida isolates from episodes of vulvovaginal candidiasis were identified by classic methodologies. The susceptibility testing of the in vitro fluconazole and ketoconazole activity against the isolates was accessed by E-test. Results: C. albicans was the most common species identified in 70% of the occurrences followed by C. glabrata (20%), C. tropicalis (7.5%), and C. guilliermondii (2.5%). In the susceptibility profile to antifungal agents, 12.5% and 16.7% of the isolates obtained from primary and episodic vulvovaginal candidiasis were resistant to fluconazole, respectively. To ketoconazole, we found that 6.25% and 12.5% of the isolates respectively from primary vulvovaginal candidiasis (PVVC) and episodic vulvovaginal candidiasis (EVVC) had high MIC values. Conclusions: E-test is a reliable method for the susceptibility testing of Candida spp. due to its simplicity, reproducibility, and lack of specialized equipment. Resistant strains and non-albicans species were verified more in cases of EVVC than in PVVC. Clinical and mycological cure of patients with episodic vulvovaginal candidiasis or complicated cases occurred after prolonged treatment and sometimes with multiple antifungals use.

Modified Slow-Strain-Rate Tensile Testing Method for Evaluation of Room-temperature Hydrogen Embrittlement Susceptibility and Its Application to 23Cr2Ni4MoV Steel

To evaluate the room-temperature hydrogen embrittlement susceptibility(HES) of high-pressure hydrogen storage vessels, a modified slow-strain-rate tensile(MSSRT) testing method was proposed for effectively aligning with their actual operating conditions. The effectiveness of the MSSRT testing method in evaluating the room-temperature HES of steels under high pressure was validated by comparing the results obtained using the conventional slow strain rate tensile(SSRT) and proposed MSSRT testing methods for 30CrMo steel, which is widely used for manufacturing high-pressure hydrogen storage vessels. The tensile properties and fracture morphologies of 23Cr2Ni4MoV steel were then examined using the MSSRT testing method under 35 MPa hydrogen and nitrogen at room temperature. Results indicate that 35 MPa hydrogen exerted a marginal effect on the tensile properties of 23Cr2Ni4MoV steel at room temperature when considering the MSSRT testing method;moreover, the test specimen basically exhibited macroscopic ductile fracture. Furthermore, obvious surface cracking was observed on the fractured specimen tested under hydrogen, whereas surface cracking was not observed on the fractured specimen tested under nitrogen. Hence, the relative reduction of area and surface cracking are necessary criteria for evaluating the room-temperature HES of steels using the MSSRT testing method. Overall, 23Cr2Ni4MoV steel might be unsuitable for manufacturing high-pressure hydrogen storage vessels.

Bile anaerobic bacteria detection and antibiotic susceptibility in patients with gallstone

OBJECTIVE: To detect bile anaerobic bacteria and antibiotic susceptibility in 59 patients with gallstoneswho had had cholecystectomy.METHODS: BACT/ALERT 120 microbe detection system and SCEPTOR microbe detection system wereused to detect bile anaerobic bacteria, antibiotic susceptibility.RESULTS: The ratio of anaerobic bacteria to the patients examined was 52.5% (31/59). Obligateanaerobe bile culture showed positive results in 4 patients. B. fragilis (37.8%) was the major type ofanaerobic bacteria in bile. Most (81.8%) of anaerobic bacteria were sensitive to metronidazole, andimipenem was suitable for β-lactamase bacteria.CONCLUSIONS: Culture of anaerobic bacteria in logarithmic phase can improve the positive rate of theculture. There are some relations between anaerobic infection and gallstone formation.

Susceptibility to Hot Cracking and Weldment Heat Treatment of Haynes 230 Superalloy

This study investigates the susceptibility of hot cracking and weldment heat treatment of Haynes 230 superalloy. The Varestriant test was conducted to evaluate this susceptibility. Welding was performed by gas tungsten arc welding （GTAW） and plasma arc welding （PAW） with stress relief heat treatment and solid solution heat treatment. A tensile test is then performed to measure the changes in the mechanical properties of the heattreated material. The results indicate that the number of thermal cycles does not affect the susceptibility of Haynes 230 superalloy to hot cracking. However, it does increase the strain. In weldment of heat treatment, stress relief annealing increases the yield strength and tensile strength of the welded parts. The section of the tensile specimens shows fibrous fractures on the welded parts, regardless of whether they are heat-treated.

EXTREMELY LOW FREQUENCY MAGNETIC FIELD SUSCEPTIBILITY OF VISUAL DISPLAY UNITS

Extremely low frequency (ELF) magnetic field susceptibility is an index of visual display unit (VDU) quality and performance. This paper provided field measured data on the susceptibility for a large variety of VDUs. A test rig was built to study the susceptibility of VDUs to magnetic fields at fundamental and third harmonic frequencies. It was found that the susceptibility level is largely dependent on refresh rate of the VDU and the orientation of the external ELF field. It was also found that the VDU susceptibility is significantly increased in the presence of harmonic frequency magnetic fields. About 30% of the tested samples have susceptibility levels higher than that stated in IEC 1000-4-8 standard.

Drug Susceptibility Patterns of Mycobacterium tuberculosis Isolates from Tuberculosis Patients in Coastal Kenya

Background: Tuberculosis (TB) is an infectious disease caused by the bacillus Mycobacterium tuberculosis. Anti-tuberculosis drug resistance is an emerging health problem in Kenya and especially in Coastal region. This is a major challenge in tuberculosis control. Diagnosis is based on Ziel-Neelsen staining alone and patients are treated without information on sensitivity patterns. Aim: This study aimed to determine drug susceptibility patterns of Mycobacterium tuberculosis in Coastal Kenya. Study Design: Hospital and laboratory based cross-sectional study was carried between April 2015 and July 2016 at Coast General Referral hospital;Tudor, Port-Reitz, Likoni Sub-County hospitals;Mlaleo, Kongowea and Mikindani health centers. Methodology: Sputum samples from patients with bacteriological confirmed TB on microscopy were cultured on Lowenstein Jensen (LJ) media. Strains of MTB complex from Lowenstein Jensen (LJ) slopes were subjected to drug susceptibility testing (DST) to first-line drugs including isoniazid (H), rifampicin (R), streptomycin (S) and Ethambutol (E) using proportional method on the Mycobacterium Growth Indicator Tube (MGIT) conventional method. Participants were offered diagnostic testing and counselling for HIV testing. Results: Drug sensitivity test was performed for a total of 210 Mycobacterium tuberculosis isolates for the first line anti-TB drugs. About seventy eight percent and twenty nine percent of the strains from new patients and previously treated patients were fully sensitive to all the drugs tested respectively. Prevalence of any resistance to one drug was 102 (48.6%, 95% CI: 20.45 - 28.23). Any single drug resistance was most frequent in isoniazid 30 (16.0%), Ethambutol 20 (10.0%), Streptomycin 18 (18.3%) and Rifampicin 4 (2.1%) in newly diagnosed patients. Among previously treated patients any resistance to streptomycin, ethambutol, isoniaziad and rifampicin was 10 (58.8%), 9 (52.9%), 7 (41.2%) and 4 (23.5%) respectively. Prevalence of MDR-TB defined as resistant to at least both isoniazid and rifampicin was 10 (4.8%) among new and previously treated patients respectively. Conclusion: The current study reveals that the overall resistance to first line anti-TB drugs was high. Although the rate of MDR-TB was relatively low, this signifies that conditions favouring the spread of MDR-TB are on high rise. Therefore, it is essential to address the problems of development of drug re-sistant strains of TB by establishing good TB programmes (DOTS). Patients’ adherence to anti-TB drugs and introducing drug sensitivity testing (DST) services at County level hospitals will minimize occurrence of drug resistant.

<i>Rhizopus oryzae</i>Retro-Orbital Abscess: Comparison of Spores and Hyphae Antifungal Susceptibility and Clinical Outcome

Rhizopus is fungus of the order Mucorales, responsible for invasive diseases called mucormycosis. The correct identification of these fungi is one of the main questions raised in the literature since the diagnosis is not simple, being the necessary detection in biopsies of infected tissues and isolation of the microorganism for identification by morphological and molecular methods. In vitro susceptibility testing results are not fully clear since there are reports showing that in vitro resistance does not always mean therapeutic failure. This paper describes a case of rhino-orbito-cerebral mucormycosis in a patient with alcoholic chronic pancreatitis and secondary mellitus diabetes involving microbiological diagnostic, antifungal susceptibility tests for conidia and hypha and clinical correlation. The causative agent was identified by morphological characteristics and DNA sequencing. Minimal Inhibitory Concentration (MIC) for amphotericin B (AMB), itraconazole (ITZ) and terbinafine (TERB) was determined by broth microdilution method. Standardization of the dynamic growth evaluation patterns using the automated system BCT (BioCell-Tracer?) was obtained allowing to determining hyphae growth control and evaluating the effect of different concentrations of amphotericin B, itraconazole and terbinafine directly on growing hyphae. The fractional inhibitory concentration (FIC) for the combination of the two antifungals that the patient received at the same time, terbinafine and itraconazole, was also determined trying to correlate with clinical outcome. Strains isolated of retro-orbital abscess were identified as Rhizopus oryzae. MIC results (MIC) testing conidia showed high sensibility to amphotericin B and itraconazole in comparison to the obtained for terbinafine. The tests in the automated system BCT using hyphae showed sensibility to all tested antifungal agents and the FIC results for combination in vitro between terbinafine and itraconazole showed synergism consistent with the therapeutic success obtained for the patient who received initial treatment with amphotericin B followed for maintenance treatment with the combination itraconazole and