Fatty Acids-Enriched Fractions of Hermetia illucens (Black Soldier Fly) Larvae Fat Can Combat MDR Pathogenic Fish Bacteria Aeromonas spp.

Aeromonas spp. cause many diseases in aquaculture habitats. Hermetia illucens (Hi) larvae were used as feed-in aquacultures and in eradicating pathogenic fish bacteria. In the present study, we applied consecutive extractions of the same biomass of BSFL fat using the acidic water-methanol solution. The major constituents of the sequential extracts (SEs) were free fatty acids (FFAs), and fatty acids derivatives as identified by gas chromatography spectrometry (GC-MS). Our improved procedure enabled gradual enrichment in the unsaturated fatty acids (USFAs) content in our SEs. The present study aimed to compare the composition and antimicrobial properties of SEs. Among actual fish pathogens, A. hydrophila and A. salmonicida demonstrated multiple drug resistance (MDR) against different recommended standard antibiotics: A. salmonicida was resistant to six, while A. hydrophila was resistant to four antibiotics from ten used in the present study. For the first time, we demonstrated the high dose-dependent antibacterial activity of each SE against Aeromonas spp., especially MDR A. salmonicida. The bacteriostatic and bactericidal (MIC/MBC) activity of SEs was significantly enhanced through the sequential extractions. The third sequential extract (AWME3) possessed the highest activity against Aeromonas spp.: inhibition zone diameters were in the range (21.47 ± 0.14-20.83 ± 0.22 mm) at a concentration of 40 mg/mL, MIC values ranged between 0.09 and 0.38 mg/mL for A. hydrophila and A. salmonicida, respectively. AWME3 MBC values recorded 0.19 and 0.38 mg/mL, while MIC50 values were 0.065 ± 0.004 and 0.22 ± 0.005 mg/mL against A. hydrophila and A. salmonicida, respectively. Thus, the larvae fat from Hermitia illucens may serve as an excellent reservoir of bioactive molecules with good capacity to eradicate the multidrug-resistant bacteria, having promising potential for practical application in the aquaculture field.

Protective potency of clove oil and its transcriptional down-regulation of Aeromonas sobria virulence genes in African catfish (Clarias gariepinus L.).

Disease episodes of fish caused by Aeromonas species are moved to the top list of limiting problems worldwide. The present study was planned to verify the in vitro antibacterial activities as well as the in vivo potential values of clove oil and ciprofloxacin against Aeromonas sobria in African catfish (Clarias gariepinus). The in vitro phenotypic virulence activities and the successful amplification of aerolysin and hemolysin genes in the precisely identified A. sobria strain were predictive for its virulence. In the in vivo assay, virulence of A. sobria strain was fully demonstrated based on constituent mRNA expression profile of tested virulence genes and typical septicemia associated with high mortalities of infected fish. Apparent lower mortality rates were correlated well with both decrescent bacterial burden and significant down-regulated transcripts of representative genes in the treated groups with clove oil, followed by ciprofloxacin as a prophylactic use for 15 days (P < 0.0001); however, the essential oil apart from ciprofloxacin significantly enhanced different hematological parameters (P < 0.05). In addition, administration of antibiotic may be considered as a pronounced stress factor in the fish even when it used in the prophylactic dose. In conclusion, medicinal plants-derived essential oils provide a virtually safer alternative to chemotherapeutics on fish, consumers and ecosystems.

Influencia de la correcta identificación en la interpretación de las pruebas de sensibilidad en aislados de Aeromonas spp. productoras de bacteriemia / The relevance of correct identification and interpretation of susceptibility testing of Aeromonasspp. bacteremia isolates

OBJETIVO: Estudiar la importancia de la correcta identificación a nivel de especie así como la interpretación de las pruebas de sensibilidad en aislados de Aeromonas spp. productoras de bacteriemia mediante los métodos convencionales rutinarios y los nuevos métodos moleculares. MATERIAL Y MÉTODOS: El estudio incluyó a 22 pacientes con bacteriemia por Aeromonas hydrophila grupo, identificadas mediante el sistema MicroScan. La identificación posterior a nivel de especie se realizó por espectrometría de masas y se confirmó mediante la secuenciación del gen rpoB. La actividad de imipenem, cefotaxima, piperacilina/tazobactam, ciprofloxacino y cotrimoxazol se estudió por microdilución comercial y tiras de gradiente de antibiótico con bajo y alto inóculo. La detección de carbapenemasas se realizó mediante el test de Hodge modificado y su confirmación mediante la detección por PCR del gen cphA. RESULTADOS: Se identificaron 9 (40,9%) aislamientos como Aeromonas hydrophila, 8 (36,4%) como Aeromonas veronii y los 5 (22,7%) restantes como Aeromonas caviae. La resistencia a los antibióticos betalactámicos mediante microdilución comercial y tiras de gradiente de CMI fue, respectivamente, del 36-50% para imipenem; del 4-56% para cefotaxima; y de 27-56% para piperacilina/tazobactam. La concordancia entre el sistema automatizado y el sistema de difusión con tira de gradiente antibiótico fue, globalmente para las 3 especies, del 68% para imipenem, del 50% para cefotaxima y del 46% para piperacilina/tazobactam. No se detectó resistencia a cotrimoxazol y ciprofloxacino por ambos métodos, aunque el 22,7% de las cepas fueron resistentes a ácido nalidíxico. CONCLUSIONES: Es fundamental la identificación a nivel de especie de los aislamientos de Aeromonas spp. ya que la resistencia a betalactámicos es especie y método dependiente. Los altos porcentajes de resistencia antibiótica encontrados no aconsejan el uso de antibióticos betalactámicos y quinolonas como tratamiento empírico de la infección invasiva por Aeromonas ssp

OBJECTIVE: To assess the relevance of correct identification and interpretation of susceptibility testing of Aeromonas spp. bacteremia isolates using newly developed molecular methods in comparison to previous conventional methods. MATERIAL AND METHODS: The study included 22 patients with bacteremia due to Aeromonas hydrophila group, microbiologically characterized using the MicroScan system. Further identification to species level was performed by mass spectrometry, and confirmed by sequencing the rpoB gene. The MIC of imipenem, cefotaxime, piperacillin-tazobactam, ciprofloxacin and cotrimoxazole was studied using a commercial broth microdilution and antibiotic gradient strips with low and high inocula. Detection of carbapenemase production was performed using the modified Hodge test, and was confirmed by amplifying the cphA gene by PCR. RESULTS: A total of 9 (40.9%) isolates were identified as Aeromonas hydrophila, 8 (36.4%) as Aeromonas veronii, and the remaining 5 (22.7%) isolates as Aeromonas caviae. Resistance to beta-lactams according to both the commercial microdilution and MIC gradient strips methods was: 36%-50% to imipenem; 4%-56% to cefotaxime, and 27%-56% to piperacillin/tazobactam. The agreement between results generated by the automated system and the diffusion antibiotic gradient strip was, for all 3 species, 68% for imipenem, 50% to cefotaxime, and 46% to piperacillin/tazobactam. No resistance to cotrimoxazole and ciprofloxacin was found by either of the two methods, although 22.7% of the strains were resistant to nalidixic acid. CONCLUSIONS: It is essential to identify the isolates of Aeromonas spp. at the species level, due to the fact that beta-lactam resistance is species- and method-dependent. The high rate of resistance to beta-lactam and quinolones reduce their application as empiric treatments for invasive infection by Aeromonas ssp

Flap infection associated with medicinal leeches in reconstructive surgery: two new drug-resistant organisms.

The use of medicinal leeches in reconstructive surgery has proven value for the salvage of flaps with venous congestion but is associated with a risk of leech-acquired infection. The most common leech-associated organism is Aeromonas hydrophila, which antibiotic prophylaxis is typically directed against. The authors describe two new multidrug-resistant organisms acquired from medicinal leech therapy that resulted in flap infection. The evaluation of suspected leech-borne infection and management protocol for this leech-acquired resistant multi-organism infection is presented.

Mortality of therapeutic fish Garra rufa caused by Aeromonas sobria.

OBJECTIVE: To investigate a case of mass mortality of Garra rufa (G. rufa) from a fish hatchery farm in Slovakia. METHODS: Causative bacterial agent was swabbing out of affected fish skin area and subsequently identified using commercial test system. Antibiotic susceptibility was determined by the disk diffusion method. RESULTS: Infected G. rufa was characterized by abnormal swimming behaviour, bleeding of skin lesions and local haemorrhages. Despite of using recommended aquatic antibiotic treatment no improvement was achieved and Aeromonas sobria (A. sobria) was identified as a causative agent of fish mortality. Due to massive fish mortality, antibiotic susceptibility of pure isolated culture of A. sobria was evaluated employing eight antibiotics against human infections. A. sobria was resistant only against one antibiotic, namely ampicilin. CONCLUSIONS: These results indicate that A. sobria can act as a primary pathogen of G. rufa and may be a potential risk factor for immunodeficient or immunoincompetent patients during the ichthyotherapy.

Enteric pathogens associated with childhood diarrhea in Tripoli-Libya.

Stool samples from children < 5 years of age with diarrhea (N = 239) were examined for enteric pathogens using a combination of culture, enzyme-immunoassay, and polymerase chain reaction methods. Pathogens were detected in 122 (51%) stool samples; single pathogens were detected in 37.2% and co-pathogens in 13.8% of samples. Norovirus, rotavirus, and diarrheagenic Escherichia coli (DEC) were the most frequently detected pathogens (15.5%, 13.4%, and 11.2%, respectively); Salmonella, adenovirus, and Aeromonas were detected less frequently (7.9%, 7.1%, and 4.2%). The most commonly detected DEC was enteroaggregative E. coli (5.4%). Resistance to ≥ 3 antimicrobials was observed in 60% (18/30) of the bacterial pathogens. Salmonella resistance to ciprofloxacin (63.1%) has become a concern. Enteric viral pathogens were the most significant causative agents of childhood diarrhea in Tripoli. Bacterial pathogens were also important contributors to pediatric diarrhea. The emergence of ciprofloxacin-resistant Salmonella represents a serious health problem that must be addressed by Libyan health authorities.

A review of necrotising fasciitis in the extremities.

OBJECTIVE: To review currently available evidence on the epidemiology and methods of management for necrotising fasciitis, with particular reference to Hong Kong. DATA SOURCES AND STUDY SELECTION: Medline, PubMed, and Cochrane Library searches of local and internationally published English language journals, from 1990 to July 2008 using the terms 'necrotising fasciitis', 'Hong Kong', 'diagnosis', 'epidemiology', 'vibrio', 'streptococci', 'clostridia', and 'management'. DATA EXTRACTION: All articles involving necrotising fasciitis in Hong Kong were included in the review. DATA SYNTHESIS: The incidence of necrotising fasciitis in Hong Kong and around the world has been increasing. This rapidly progressive infection is a major cause of concern, due to its high morbidity and mortality. Up to 93% of affected patients at our hospital were admitted to the Intensive Care Unit and many still died from septic complications, such as pneumonia and multi-organ failure. Radical debridements in the form of amputations and disarticulations were considered vital in 46% of the patients. Early recognition and treatment remain the most important factors influencing survival. Yet, early diagnosis of the condition is difficult due to its similarities with many other soft tissue disorders such as cellulitis. Repeated surgical debridement or incisional drainage continues to be essential for the survival of sufferers from necrotising fasciitis. Many authorities have reported that carrying out the first fasciotomy and radical debridement within 24 hours of symptom onset was associated with significantly improved survival, which also emphasises the importance of early diagnosis. CONCLUSION: Clinicians must adopt a high index of suspicion for necrotising fasciitis. Empirical antibiotics must be started early and repeated physical examinations should be performed, while maintaining a low threshold for tissue biopsy and surgery. The timing of the first fasciotomy and radical debridement within a window of 24 hours from symptom onset is associated with significantly improved survival.

Immunization with bacterial antigens: furunculosis.

Although the nature of the antigens and the immune responses they elicit to achieve immunity to furunculosis are still not well defined, the currently available vaccines comprising A. salmonicida bacterins emulsified in oil adjuvants and delivered by intraperitoneal injection provide remarkably high levels of long-lasting protection. Despite some concern over side-effects, these vaccines have been adopted by most Atlantic salmon farmers over the last four years, transforming a situation where furunculosis outbreaks were becoming catastrophic to one where losses from the disease are negligible. Present evidence indicates that antibody responses to the polysaccharide capsule and iron regulated outer membrane proteins are associated with protection. Furthermore, cell-mediated immune responses involving antigen-induced release of cytokines from lymphocytes and the resultant activation of macrophages with the ability to kill the pathogen are also considered important protective mechanisms. Vaccines comprising whole A. salmonicida cultures grown under iron-restricted conditions and delivered by injection in an oil adjuvant are expected to induce prolonged stimulation of all the above responses. While these vaccines are suitable and effective for administration to salmon smolts there is still a need for mass vaccination by immersion or oral routes for salmonid fry. Effective means of achieving this are still required.