Mouse embryo development under an enriched atmosphere of negative air ions: a pilot study

Several studies have shown that negative air ions may have positive effects on the living organisms. The aim of our study was to evaluate whether an enriched environment of negative air ions have an influence on the mouse embryo development. A controlled negative air-ion generator (CNAIG) was installed in a standard incubator and test embryos cultured under the enriched atmosphere with negative ions. Embryo development was judged in terms of blastocyst rates, blastocyst total cell number, and potential of these to develop to term. In a first series of experiments, we observed that mouse embryos cultured in the atmosphere enriched with negative ions developed up to the blastocyst stage with a 96.1% (49/51) efficiency, similarly (p= 0.675)to controls 97.1% (32/34). Blastocysts obtained in the experimental group showed a total mean number of cells (110±20.1) statistically equivalent (p = 0.351) to controls (121±33.4) cultured under standard conditions. In a second series of experiments, blastocysts were transferred to synchronized pseudo-pregnant females, so embryo development could be followed in vivo. A total of 33.3% of the blastocysts transferred from the experimental group developed to term, in a similar proportion (p>0.05) to the control group (40%).In this pilot study, we demonstrate for the first time that mouse embryos cultured in an enriched negative ions atmosphere are able to develop in vitro and in vivo with very high successful rates. Key words: controlled negative air-ion generator (CNAIG), mouse embryo development, in vitro fertilization, embryo culture

Screening of Chlamydia trachomatis urogenital infections among the male and female population of the Republic of Macedonia

Noninvasive urine screening for Chlamydia trachomatis infections offers a valuable public health tool, that could be of vast importance in Chlamydia control programs. The goal was to determine the prevalence of C. trachomatis infections among a sexually active population, to define the epidemiological factors associated with it, and to develop potential selective screening strategies among asymptomatic individuals in the Republic of Macedonia, using a highly sensitive and specific DNA amplification method for C. trachomatis. A total of 1435 urine samples, divided into two main groups: asymptomatic individuals (n = 1210) and symptomatic patients (n = 225), were tested. Samples from the asymptomatic group were collected during routine screening programs, while the symptomatic group consisted of patients with symptoms of urogenital tract infection, attending sexually transmitted diseases (STD) clinics. The presence of C. trachomatis was determined using commercial AMPLICOR C. trachomatis Assay (Roche Diagnostic Systems, Inc., Branchburg, NJ, USA). The prevalence of C. trachomatis infections among different groups was: recruits 0%, soldiers 0.4%, policemen 3.5%, clerks 4.6%, pregnant women 4%, and students 4.4%. The average prevalence for both groups (asymptomatic and symptomatic) was 2.3%[95% confidence interval (CI): 1.5-3.1%]. The average prevalence for the asymptomatic group was 1.6% (95% CI: 0.8-2.4%), while the average prevalence for the symptomatic group was 6.2% (95% CI: 3.1-9.3%) which were significantly different (P = 0.00003). Testing first void urine specimens by AMPLICOR C. trachomatis assay is a highly sensitive and specific method for diagnosing C. trachomatis infections in men and women. This method provides health care workers and public health officials with a new molecular amplification assay that uses noninvasive urine specimens for population-based screening purposes. The prevalence of C. trachomatis was relatively low among asymptomatic individuals. However, selective screening strategies are highly recommended for testing the student population in the Republic of Macedonia. Journal of the European Academy of Dermatology and Venereology 08/2005; 19(4):427-30. • 2.69 Impact Facto

Perspectives for applications of nanotechnology and nanomaterials in medicine

Nanotechnology is a new science that comprise several science fields (chemistry, biology, physics, material science and engineering) conducted devices or materials that have at least one dimension undersize of 100 nm. Nanomaterials are chemically or physically different to their macro-scale counterparts and may have unique health and environmental impacts. Nanoparticles can be synthesized by reactions in solid state (breaking the larger materials) and chemical methods (wet chemical synthesis). Extremely small size enables nanoparticles to enter the human body through usual or unusual routes, pass through cell membranes, or cross the blood-brain barrier. At present, potential use of nanotechnology in a range of applications at any stage in food industry: production, processing, packaging, labelling, transporting, tracing, keeping the quality of food product and extend the product shelf-life, leading to less food waste, water filtration, removal of undesirable tastes, flavors or allergens from food products. Applications of nanotechnology and nanomaterials in medicine (Nanomedicine) based on Nano (bio) sensors enables rapid and sensitive detection of pathogenic bacteria and trace levels of viruses in small sample volumes, at lower costs. Early detection of pathogens enables accurate and prompt treatment. Sensors to detect biofilm formation by bacteria on surfaces are developing. Multi-drug resistant microorganisms (MDR) are one of the most serious and increasing global, public health threat. New strategies urgently needed to combat MDR, includes nanomaterials as very promising approach. Metal nanomaterials (silver, gold, copper, titanium, zinc, magnesium, cadmium, and alumina) possess unique antimicrobial activities. Researchers are developing: - Method to release insulin from sponge-like matrix and Nano-capsules, when the glucose level rise - Nano-particles to be taken orally (pills), could pass the intestines into the bloodstream - Sensors to detect bacterial biofilms on surfaces - Nano-particles to defeat viruses by delivering an enzyme that prevents their reproduction in the patients bloodstream - Gelatin nanoparticles can be used to deliver drugs to damaged brain tissue - Nanoparticles to deliver vaccine, allowing the vaccine a stronger immune response Risks of nanotechnology are still unknown and unpredictable. Keywords: applications, nanomedicine, nanoparticles, nanotechnolog

Hepatitis B and hepatitis C viral infections, importance, microbiological diagnosis and incidence in Shtip, Republic of Macedonia

Liver inflamation may be caused by various factors. Infectious agents as most important factors includes: viruses, bacteria and parasites. Most frequent infective hepatitis worldwide are viruses: Hepatitis A, B, C, D and E. Diagnosis is performed on the basis of symptoms and signs of the patients, anamnesis, laboratory testing, CT and MR and liver biopsy. \ud Laboratory (serologic) tests includes biochemical tests of blood, detection of enzymes (ALT, AST), detecting of Hepatitis B markers (antigens: HBs, HBc, HBe and antibodies: anti HBs, anti HBc and anti HBe), and fro Hepatitis C: anti HCV antibodies. Detecting of viral DNA for Hepatitis B and viral RNA for Hepatitis C is of great importance, using PCR (Polymerase Chain Reaction). During period from 2010 to 2015 година, in municipality of Sthip, Republic of Macedonia, from total examined 5149 persons, positive results were for Hepatitis B infections in 171 (3,3 %), from which 97 (56,7 %) male and 74 (43,3 %) female persons. During same period, from a total of 3253 examined persons for Hepatitis C infections, 56 (1, 7 %) had positive results. From 38 registered addicts three were positive for Hepatitis B (8, 6 %), 32 for Hepatitis C (91, 4 %), and three positive for both (Hepatitis B+C). From 350 prisoners of prison in Sthip, 35 (10%) were positive for Hepatitis C. In the Centre for dialysis in Sthip, from 51 patients, 18 were detected positive for Hepatitis B and 18 positive for Hepatitis C and 15 negative for both infections. Key words: Hepatitis B, Hepatitis C, diagnosis, incidence, treatment

The influence of reactive oxygen species on in vitro fertilization success

The reactive oxygen species (ROS) are one of the serious factors which negatively affect the in vitro fertilization (IVF) outcome. When the formation of ROS overcome the ability of the biological system to detoxify them, they cause damage of the cells known as oxidative stress. These oxygen derived free radicals normally are produced from mitochondria during normal oxidative respiration. Due to their highly reactive nature, ROS can combine directly with other molecules leading to their structural and functional changes which result in cellular damage. Physiologically ROS can modulate the reproductive processes such as sperm-oocyte interaction, early embryo development and implantation. However the imbalance in their presence can cause oxidative stress affecting the pregnancy outcome. Although the IVF methods are continuously developing and improving there are still not sufficient microenvironment conditions that will avoid ROS accumulation and their negative influence on IVF outcome. The source of ROS during IVF may be due to the absence of endogenous defense mechanisms or due to the use of various manipulative techniques. In that way the ROS can originate from the endogenous production from the gametes and embryos or can be the result of the external factors such as: culturing media, O2 concentration, light or the manipulation technique which has been used. One of the most important endogenous factor is increased oxidative stress in the spermatozoa which is associated with poor fertilization rate, low embryo quality and high rate of pregnancy loss. The other endogenous factors are connected with oocytes and the follicular fluid. It has been shown that higher ROS levels can trigger disruption of the oocyte cytoskeleton, affect the spindle formation, aneuploidy and embryo developmental arrest. Since embryo is a fast developing organism that needs high energy in some cases can also generate excessive amounts of ROS. Pathological levels of ROS during embryo culturing result in a low quality embryos with high fragmentation leading to low clinical pregnancy rates. To avoid the effects of the known external sources of ROS many improvements were made by additions of supplements in the culture media that reduce the ROS formation, culturing under lower O2 concentrations, reduced exposure to light and introduction of vitrification methods for cryopreservation. Although many strategies are implemented so far, there is still a need for further development of new ways to avoid the oxidative stress as much as possible during the IVF treatments. One of the future approaches to prevent the harmful oxidative stress on IVF may be optimization of the environment by using the enriched atmosphere with negative air ions. Key words: IVF, pregnancy, ROS, fertilizatio

Aflatoxins and ochratoxins, harmful mycotoxins for human and animal health

Aflatoxins and ochratoxins are toxic and carcinogen mycotoxins produced as secondary metabolites by several moulds species. Aflatoxins are producing by Aspergillus flavus and Aspergillus parasiticus in dry conditions and high temperatures. Food and feed may be contaminated during harvest, transport, and storage and processing. Contaminated food, imported in human body may cause harmful effects including liver cancer. The aflatoxins consist of about 20 similar compounds but only four (B1, B2, G1 and G2) are naturally found in foods. Aflatoxin B1 is the most commonly found in food and also the most toxic. If lactating animals ingest aflatoxins by contaminated feed, toxic metabolites may be present in milk. Aflatoxin M1 and M2, are important contaminants in dairy products. Cereals, nuts, figs, tobacco and many other food products may be contaminated by aflatoxins. The EU sets limits for aflatoxin B1 and for total aflatoxins (B1, B2, G1 and G2) in nuts, dried fruits, cereals and spices range from 2-12 μg/kg for B1, from 4-15 μg/kg for total aflatoxins and 0.050 μg/kg for aflatoxin M1 in milk and milk products. Limits of 0.10 μg/kg for B1 and 0.025 μg/kg for M1 have been set for infant foods. US food safety regulations include a limit of 20 μg/kg for total aflatoxins (B1, B2, G1 and G2) in all foods and 0.5 μg/kg for M1 in milk. Higher limits apply in animal feeds. Ochratoxins are secondary metabolites produced by Aspergillus ochraceus, Aspergillus niger and Penicilinum verrucosum. Ochratoxins includes: ochratoxin A (OTA), B, C, α and β. The most important and most toxic, found naturally in food, is ochratoxin A (OTA). The ochratoxin B, is rare and much less toxic, also may be found in food. Ochratoxins may be detect on various types of food and feed, in conditions of higher moisture and high temperatures, during harvest transport, storage and processing. Ochratoxins are toxic for kidneys and liver. Because of toxic harmful actions for human and animal health, food and feed protection aflatoxins and ochratoxins is very important. Separation of contaminated from non-contaminated food and feed is essential for prevention of human and animal health and high economy savings. Real situation with contamination of food and toxic effects of aflatoxins and ochratoxins on human and animal health are unknown. Key words: aflatoxin, Aspergillus, ochratoxin, secondary metabolite

Importance of probiotics for human health

The gut flora is a complex community of about 1000 different types of microorganisms (bacteria, yeasts and viruses) of which the greatest majority are bacteria. Most of them lives in colon. Bacteria of gut flora belongs to group of beneficial („good“) microorganisms, but also bacteria that cause different infections and diseases („bad“ microorganisms) may be present. Beneficial microorganisms suppress growth of bad bacteria and yeasts. Gut flora has very important role for human health. It produces vitamins (vitamin K and some vitamins of B group), also stimulates the immune system and regulates integrity of gut. Balance changes of gut flora are reasons for numerous diseases. Probiotics are live bacteria belonging to the group of „good“ bacteria. They can stop pathogenic bacteria by competition, growth inhibition and attaching to gut epithelium. Numerious studies have proved many beneficial activities of probiotic use on immune system, suppression of pathogenic bacteria, gastrointestinal diseases, post antibiotic diarrhea, hypercholesterinemia, colorectal carcinoma, inflammations, depressions and anxiety, high blood pressure and some skin disorders. Influence of probiotics to human health is a reason for wide use in all age groups. Investigation and analyzes of several health institutions in Sthip, city in Republic of Macedonia, showed that importance of probiotic for promotion of human health is very well known to medical personal and most appropriate probiotics for specific indications are recommended. Most recommended probiotics are: Linex, Probalans, Prolife, Darmflora plus, Helicobalans, Biogaja and Diastop. The best natural sources of probiotics are yogurt, ayran and kefir. Key words: probiotics, gut flora, Lactobacillus, Bifidobacteriu

Brucellosis - re-emerging zoonotic and food borne disease

Introduction Zoonotic diseases primarily affects domestic and wild animals than transfer to humans. Depending on the agent, way of spreading may be respiratory, contact, alimentary (food or water) or combination. Brucellosis is a re-emerging zoonotic disease which is spreading by all mentioned ways. Disease exists in R. Macedonia since 1980 with over 12.000 reported and confirmed human cases. Disease is reported in all neighbouring and almost all European countries with significantly different incidence. Aim To present current epidemiology situation, ways of spreading of the disease in respect of zoonotic and foodborne disease, and measures for control and prevention of brucellosis in Republic of Macedonia, a small country with endemic areas, long history and experience. Material and Methods Review and presentation of official data on epidemiology of brucellosis in past 36 years and situation after implementation of new national strategy based on vaccination of small ruminants. Results Brucella melitensis biotype 2 was confirmed as etiological agent in R. Macedonia. Recent study based on molecular methods for species typing (AMOS PCR and RT PCR), and genotyping (MLVA-16 and MLVA-8), beside Brucella melitenisis also confirmed Brucella abortus (for the first time in Macedonia). Epidemiological data suggested about 23% of spreading the disease by alimentary way (foodborne disease due to consumption of unpasteurized milk, cheese, and undercooked infected meat), 34% by contact and 43 % by combined way of spreading brucellosis. Respiratory way is not often, happens in laboratories or working with infected animals. About 80% of patients lived in rural and 20% in urban areas. Disease in Mediterranean area has seasonal character with maximum in May-June and minimum in winter. Since 2008, in R. Macedonia, national control strategy was completely changed from „test and slaughter” to vaccination of small ruminants (sheep and goats) with Rev 1 vaccine, applied intraocular. Results are significant decreasing of epizooty in animals and human morbidity (287, 167, 107, 94, 47, 35, 20 and 23 in 2009, 2010, 2011, 2012, 2013, 2014, 2015 and 2016 respectively). Conclusions Control of Brucellosis is very complicated due large reservoirs in domestic and wild animals. Control of animal brucellosis is imperative for control of human brucellosis. Preventive measures includes effective veterinary and health control of animals (trading and transport and slaughter) and animal products (meat, milk and their products), education of the population, continuously state financial support, institutional cooperation and regional cooperation. Key words: brucellosis, control, epidemiology, foodborne

Mouse embryo development under an enriched atmosphere of negative air ions: a pilot study

Several studies have shown that negative air ions may have positive effects on the living organisms. The aim of our study was to evaluate whether an enriched environment of negative air ions have an influence on the mouse embryo development. A controlled negative air-ion generator (CNAIG) was installed in a standard incubator and test embryos cultured under the enriched atmosphere with negative ions. Embryo development was judged in terms of blastocyst rates, blastocyst total cell number, and potential of these to develop to term. In a first series of experiments, we observed that mouse embryos cultured in the atmosphere enriched with negative ions developed up to the blastocyst stage with a 96.1% (49/51) efficiency, similarly (p= 0.675)to controls 97.1% (32/34). Blastocysts obtained in the experimental group showed a total mean number of cells (110±20.1) statistically equivalent (p = 0.351) to controls (121±33.4) cultured under standard conditions. In a second series of experiments, blastocysts were transferred to synchronized pseudo-pregnant females, so embryo development could be followed in vivo. A total of 33.3% of the blastocysts transferred from the experimental group developed to term, in a similar proportion (p>0.05) to the control group (40%).In this pilot study, we demonstrate for the first time that mouse embryos cultured in an enriched negative ions atmosphere are able to develop in vitro and in vivo with very high successful rates. Key words: controlled negative air-ion generator (CNAIG), mouse embryo development, in vitro fertilization, embryo culture

An Update on Brucellosis: Endemic and Potential Global re-Emerging Zoonotic and Foodborne Disease

Brucellosis is a re-emerging zoonotic disease, which spreads in different ways: respiratory (inhalation), contact, alimentary (consumption of unpasteurized milk and contaminated dairy products) or a combination of these. The disease has existed in the Republic of Macedonia since 1980, with over 12.000 reported and confirmed human cases. All neighbouring and many other European countries have also reported existence of brucellosis with significantly different incidence. Brucellosis remains a rare disease in EU/EEA. In 2014, 354 confirmed cases of brucellosis were reported by 18 EU/EEA countries. The highest rates were reported by Greece (135), Spain (60) and Portugal (50). The control of brucellosis is very complicated due to large reservoirs in domestic and wild animals. The control of animal brucellosis is imperative for the control of human brucellosis. Key words: brucella, brucellosis, food-borne, re-emerging, reservoirs, zoonotic