In hospital environments, various factors such as temperature, relative humidity, hospital building design, ventilation system, and indoor population density and disinfection methods can affect the concentration of air pollutants (Mousavi et al. 2019). In this study, a total of 262 samples of bacteria and fungi from 10 hospital wards were examined. As a result, 14 bacterial species and 12 fungal species were identified.
Particle distribution in different wards of the hospital
Particle density in different wards of the hospital is influenced by factors such as the number of beds in each room, the number of patients in the hospital room, the rate of ventilation, the number of staff and the proximity to a street (Shokri et al. 2016). Due to the effect of particles on the density of bacterial and fungal bioaerosols, the sampling time of suspended particles coincided with the fungal and bacterial sampling. The results of this study showed that the average particle concentration in about 90% of the hospital wards and ambient air was higher than the 24-hour World Health Organization (WHO) and United States Environmental Protection Agency (USEPA) standards, which are and, respectively. In accordance with our observations, Rezaei et al. reported the concentration of PM10 and PM 2.5 suspended particles in the air of some wards inside the air around a hospital in Tehran above the recommended limits of the WHO and USEPA and stated that the quality of indoor air was affected by the ambient air (Rezaei et al. 2013).
In this study, the effect of temperature as one of the meteorological parameters on the concentration of suspended particles was investigated. Based on the results, the average temperature of the hospital wards was 24.77°C. Centers for Disease Control and Prevention (CDC) and Healthcare Infection Control Practices Advisory Committee (HICPAC) recommend temperatures of 21–24°C and 23–27°C, respectively, in winter and summer for most wards of hospitals (Chinn &Sehulster 2003).
In the infectious part, in all concentrations studied, temperature and particle concentration had a significant relationship and a positive correlation coefficient, i.e. with increasing temperature in the infectious part, the concentration of suspended particles increased. This increase in the concentration of particles in the ICU and infectious wards can be due to reasons such as the large number of patients admitted to the ward, high traffic of companions, use of non-sterile personal equipment by companions and patients, lack of proper ventilation system, and type of disease. Therefore, temperature can be considered as a factor in increasing the concentration of particles. Shokri et al. reported a significant positive relationship between the temperature and humidity and the concentration of suspended particles in the sizes of 0.3, 2.5 and 10 µm in indoor air (Shokri et al. 2016).
According to the results, there was no direct and significant relationship between moisture and particle concentrations in different diameters in all the wards. The most polluted part in terms of the presence of particles in different sizes was related to the emergency ward for reasons such as a wide range of patients and their congestion in the emergency ward compared to other wards or high traffic of clients, lack of hygiene, smoking and inadequate ventilation. Unlike, it was found that the operating room befitted from the best conditions, which can be due to the high level of health standards such as limited traffic, fewer patients, closed entrance to the other wards and proper ventilation, sterilization and disinfection of surfaces and premises. In agreement with our study, Nikpei et al. reported that the highest number of particles with diameters of 0.3 and 2.5 µm was for the emergency ward (Shokri et al. 2016).
Fungi
According to the data from Table 1, the lung ward was the most infected part of the hospital in terms of the presence of airborne fungi. Even so, in this section, the number of colonies was higher than 200 CFU/m3. Also, the operating room with the lowest number of grown colonies had the best air quality among the other wards of the hospital. The low level of pollution in the operating room can be due to the importance and sensitivity of this ward compared to other wards inside. The hospital has a high level of compliance with health standards such as limited traffic, fewer patients, frequent purification of room air by high-efficiency particulate air (HEPA) filters, suction of outside air by exhaust fan and sterilization of equipment and surfaces by ultraviolet devices.
In this study, the number of colonies measured in different wards of the hospital was higher than the standard recommended by the WHO, which is 50 colonies. Although the health hazards posed by bioaerosols have been identified and proven, no specific permissible limits are recommended for this category of airborne pollutants and the values provided are the recommended values. The large number of hospital beds and the consequent increase in the number of patients and visitors, as well as the inadequate ventilation system in some patients' rooms and the poor quality of indoor air, are among the factors providing conditions for growth and proliferation of fungal spores (Azizifar et al. 2009, Kamali Sarwestani et al. 2017). Some clients' activities such as talking, walking in the wards, sneezing and coughing consecutively also increase the spread and consequently cause the load of bioaerosols in the air of the hospitals to increase (Hoseinzadeh et al. 2013). Azizifar et al. reported that the average concentration of fungi in different wards of the hospital was 200 CFU/m3. In the current study, the highest and lowest infections were related to the infectious ward and operating room (ear, nose, throat and eyes) with the average concentrations of 300 and 94 CFU/m3, respectively (Azizifar et al. 2009).
Among the fungal species observed in the study hospital, Penicillium, Cladosporium, Aspergillus Niger and Aspergillus Flavus were the most abundant, respectively. Similarly, Cladosporium, Penicillium and Aspergillus have been reported as the dominant species isolated from hospital environments. For example, Azizifar et al., who quantitatively and qualitatively evaluated fungal air pollution in different wards of the hospital, stated that the highest percentage of fungi observed in the hospital air were Penicillium (36.36%) and Cladosporium (24.74%), and Aspergillus niger (17.97%) (Azizifar et al. 2009).
Table 1
Mean and standard deviation of fungal pollution load (CFU/m3) in the hospital
Ward
|
Internal men
|
Internal women
|
Lung
|
Neurology
|
Infectious
|
Burn
|
ICU
|
Operating room
|
Emergency
|
Outside air
|
Average (CFU/m3)
|
62.20
|
124.25
|
223.09
|
154.09
|
63.27
|
42.70
|
55.55
|
0.42
|
106.42
|
52.73
|
Standard Deviation (CFU/m3)
|
26.72
|
283.79
|
164.98
|
25.38
|
12.66
|
24.07
|
13.03
|
0.50
|
71.23
|
27.51
|
The reason for the predominant presence of these three types of fungal species in the air inside the hospital can be stated that Penicillium, Cladosporium and Aspergillus fungi have high growth ability in different climatic conditions and by producing small, light spores remain in air. Conidia spores of these fungi also have an outer layer rich in hydrophobic protein, which leads to their further suspension in the air. These fungi have the ability to supply the carbon and hydrogen they need from a variety of sources, allowing them to survive longer in a variety of conditions. However, other fungi such as Alternaria and Ulocladium and some other fungi produce smaller, larger and heavier spores that tend to settle faster and are found at different levels (Alangaden 2011, Vonberg &Gastmeier 2006).
The most and least fungal species observed in the present study were Penicillium and cranosporium and yeasts and scopolariopsis (Fig. 1). Several species of fungi have been observed in some stations. Fungal species of Aspergillus flavus, Alternaria, Penicillium and Geotrichum were mostly observed in the burn ward. In the ICU ward, Alternaria, Aspergillus niger and Ulocladium were mostly observed. Furthermore, in the emergency ward, the most observed species were Cladosporium, Penicillium, Rhizopus and Ulucladium.
Bacteria
Bacterial densities in the hospital wards ranged from 3.75 to 214.2 CFU/m3. According to the results presented in Table 2, men’s ward and operating room with an average of 148 ± 65 and 14.83 ± 16.05 CFU/m3 had the highest and lowest levels of contamination load. The main reason can be considered the observance of high level of health standards in the operating room compared to other hospital wards, as well as disinfection and proper ventilation.
It should be noted that the average density of bacterial bioaerosols in the men’s and women’s wards, lungs and burns was higher than the standard recommended by the WHO (100 CFU/m3) (Fig. 2). According to the study by Valedeyni Asl et al., the density of bacterial bioaerosols in the air of Ardabil teaching hospitals was higher than the standards suggested by the WHO and USEPA. Factors such as population density, ventilation and health conditions of the building and the type of hospitalized patients, the presence of companions and staff can increase the density of bacteria compared to the proposed standard (VALEDEYNI et al. 2018).
Table 2
The mean of bacterial number observed in each ward
Ward
|
Average
|
Standard Deviation
|
p
|
Internal men
|
148.00
|
65.154
|
0.071
|
Internal women
|
142.00
|
54.513
|
Lung
|
113.58
|
37.378
|
Neurology
|
92.66
|
28.053
|
Burn
|
133.50
|
39.877
|
ICU
|
90.33
|
18.712
|
Infectious
|
100.08
|
99.235
|
Operating room
|
14.83
|
16.059
|
Emergency
|
38.91
|
20.887
|
Outside air
|
84.50
|
59.046
|
Considering that Tohid Hospital in Sanandaj is an educational and medical center, it can be said that one of the reasons for the high density of bacterial bioaerosols in this hospital, in addition to the high volume of patients, can be the presence of many students. In Masoudinejad's study, a significant linear correlation was observed between patients and population density with the concentration of bacteria, which showed the larger the population, the higher the number of bacteria in the air (Massoudinejad et al. 2015).
Environmental parameters are one of the factors affecting the microbial population in hospital environments. Due to the fact that temperature changes in the studied areas are in the small range (24 to 25), it does not have a significant effect on the concentration of bacteria. Also, the lack of correlation between the percentage of relative humidity and the number of bacteria in this study can be attributed to the small range of relative humidity changes (23–27) at the sampling points, which is 40 to 60% less than the proposed standard (Rafiee et al. 2018).
Since only floors, walls and some pieces of equipment are washed with disinfectants when washing and disinfecting parts, the humidity of the air can increase due to washing, which facilitates the growth and persistence of microorganisms. On the other hand, if air conditioning is used in hospitals, it can reduce the humidity; because humidity is effective in the growth of bioaerosols and if air conditioning is used, bioaerosols that are in the outside air do not penetrate into the indoor environment and therefore the amount of indoor air pollution will be reduced. As it turns out, different findings have been presented regarding the effect of temperature and humidity on the growth of bacteria in the air of hospitals, which requires more and more detailed studies in this field.
It was found that there was no significant relationship between the number of particles observed in different sizes and concentrations and the number of bacteria observed. That is, particle concentrations and different particle sizes had no effect on the microbial load. The findings of this study are inconsistent with some previous studies. Mirhosseini et al. reported that there was a significant relationship between 1 to 5 µm particles and the density of bacterial bioaerosols in the surgical and ICU wards (Mirhoseini et al. 2015).
According to the results of differential tests, most of the bacteria isolated from the air of the wards in Tohid Hospital was Staphylococcus hemolyticus (Fig. 3). In a study by Solomon et al., performed in Ethiopia, Staphylococcus coagulase-negative bacteria, Staphylococcus aureus, Enterococcus faecalis, Enterococcus faecium, Acinetobacter, Escherichia coli, Pseudomonas aeruginosa were detected in a university teaching referral hospital (Solomon et al. 2017).
Staphylococci are among the opportunistic microorganisms that are detected in most areas, and since Staphylococcus species are part of the natural flora of the skin and nose, it seems that their high rate in this study due to the increase in population, especially during the presence of companions. Coagulase-negative staphylococci include species of the genus Staphylococcus that lack the coagulase enzyme, and the most important species are Staphylococcus epidermidis, Staphylococcus saprophyticus, and Staphylococcus hemolyticus. These species are not highly toxic, but are an important cause of infections in high-risk groups.
Staphylococcal infections can be transmitted through contact with an infected person or the patient's belongings, including clothing, towels, and bedding. The staff of hospitals are the most important and common carriers of them, from one sick person to another (DADASHI et al. 2015). Staphylococcus hemolyticus is the second most common coagulase-negative staphylococcus as a pathogen in normal valvular endocarditis, septicemia, bacteremia, bone and joint infections, urinary tract infections, wound infections, and opportunistic infections. Unlike other CoNS, Staphylococcus hemolyticus is resistant to vancomycin (Sivagnanasundaram et al. 2019).
Bacillus was one of the bacteria identified in the hospital wards. The ability to form spores enables Bacillus species to tolerate harsh environmental conditions and conventional disinfection methods. Some bacilli are part of the opportunistic natural flora of the human gastrointestinal tract and are dangerous to immunocompromised patients such as those with AIDS and diabetes. Micrococcus luteus is also known as a nosocomial pathogen and causes pneumonia, endocarditis, meningitis and intracranial hemorrhage (Naddafi et al. 2009, Sivagnanasundaram et al. 2019).
In this study, Escherichia coli was less common than Staphylococcus, Bacillus and Pseudomonas. It has been documented that Escherichia coli is involved in the development of diseases such as urinary tract infections, sepsis, pneumonia, gastroenteritis and meningitis (Sivagnanasundaram et al. 2019).
According to the results, the total average of bioaerosols in this hospital was 184.38 CFU/m3. The highest density of bioaerosols was in the lung ward with the amount of contamination (336.67 CFU/m3) and the lowest was in the operating ward with the amount of contamination (15.25 CFU/ m3). Following that, the women’s ward and then neurology had the highest pollutant density.
Evaluation of bacterial resistance to antibiotics
The results of antibiotic resistance of 18 common bacteria detected in this study have been presented in Table 3. Among them, the highest drug resistance was observed in Staphylococcus hemolyticus and aureus. Staphylococcus hemolyticus had shown resistance to five types of GM-CP-AZM-AMX-CFM antibiotics.
Table 3
Antibiotic-resistant bacteria
Type of bacteria
|
Antibiotic resistant
|
Staphylococcus hemolyticus
|
GM- CP- AZM- AMX- CFM
|
Staphylococcus aureus
|
GM - AMX- CFM
|
Pneumococcus
|
SXT- CFM- GM
|
Staphylococcus CONs (epidermidis)
|
CP- GM
|
Staphylococcus CONs (Saprophyticus) and Staphylococcus CONs (Epidermidis) and Bacillus
|
CFM
|
Lateus micrococcus
|
AMX
|
Streptococcus pyogenes
|
GM
|
Staphylococcus aureus is a ubiquitous organism and has a high potential for causing various diseases in humans due to its high resistance to adverse environmental conditions. The development of resistance to various antibiotics in strains of Staphylococcus aureus causes many problems in the treatment of diseases related to these microorganisms, thus, it is necessary to know the pattern of resistance of these microorganisms in the treatment of related diseases (Saha et al. 2008). Antibiotic resistance of Staphylococcus aureus to GM-AMX-CFM antibiotics has been observed. In the study by Saadat et al., the highest resistance of Staphylococcus aureus to amoxicillin was found, which is close to the findings of this study (Saadat et al. 2014). Also, Staphylococcus epidermis was resistant to CP, GM and CFM. In the study by Nourbakhsh and Momtaz, Staphylococcus epidermis was the most resistant to the antibiotics erythromycin, ciprofloxacin, clindamycin and tetracycline, which is consistent with the results obtained in the present study (Nourbakhsh &Momtaz 2016). In the study by Dadashi et al., on the frequency and profile of antibiotic resistance of Staphylococcus coagulase negative specimens in the medical staff of an army hospital, 62% of the isolates were CNS strains (Staphylococcus negative coagulase). The highest resistance in these strains (epidermidis, saprophyticus, hemolyticus and hominis) against oxacillin: 90, 94, 89 and 100%, and gentamicin: 86, 81, 65 and 100%, respectively, and the highest susceptibility was observed in mupirocin (100, 100, 100 and 100%), vancomycin (85, 88, 78 and 100%) (DADASHI et al. 2015).
According to the findings of this study, the bacteria were most resistant to the antibiotic gentamicin, so that about 70% of the bacteria tested for antibiotic resistance were resistant to gentamicin. After that, the highest resistance of bacteria was to cefixime, amoxicillin and ciprofloxacin, respectively. Based on the results of this study, while identifying the underlying factors and appropriate measures to control antibiotic-resistant bacteria, it is recommended for coagulase-negative strains of Staphylococcus, especially species such as Staphylococcus hemolyticus to avoid prescribing high-resistance antibiotics such as gentamicin, cefixime, amoxicillin, and ciprofloxacin to these strains.
The PCR assay was performed for two samples of the bacteria that had the highest resistance and the highest presence among the identified bacteria. Sample number one, resistant to five types of antibiotics (ciprofloxacin, gentamicin, azithromycin, amoxicillin, cefixime) and sample number two was resistant to four types of antibiotics (gentamicin, azithromycin, amoxicillin, cefixime). The results of molecular PCR test for both samples showed Staphylococcus hemolyticus. The reason for selecting these two samples was the highest resistance to antibiotic discs, the highest frequency and repetition among the samples taken during the study, as well as different appearance characteristics. NCBI nucleotide BLAST search (http://blast.ncbi.nlm.nih.gov/Blast.cgi) showed that sequences share highest identity with Staphylococcus haemolyticus, Bacillus subtilis, Bacillus licheniformis (Fig. 5).
Modeling the dispersion of pollutants using SURFER software
Maps prepared based on data from particle measuring stations in lung, neurology, infectious and open air wards showed that the emission and concentration of pollutants in the eastern parts had a higher dispersion. In other areas, including burns, ICU, emergency and men’s and women’s wards, the distribution of pollutants in all measuring stations in these areas was high. According to the results, the entrance of the infectious ward (east side) has the highest particle density, which can be due to the location of the waiting room at the entrance of the ward and high traffic in these areas (Fig. 6). However, the distribution of pollutants in other wards was evenly distributed.
Counter curves on the maps related to the distribution of bacteria in the hospital wards showed that in the men’s and women’s wards, the concentration of bacteria in the stations located in the northern and central parts of the wards was higher than the southern parts. The number of bacteria in the operating room was measured in the range of 2–36 CFU/m3. The counter curve numbers show that the bacterial dispersion was lower in most parts of the measuring stations and the less contour lines in the section. The central part shows the high concentration of bacteria. In the previous sections, it was stated that the lowest bacterial density was in the operating room.
In the intensive care unit, bacterial densities were measured in the range of 72–128 CFU/m3, and counter curves in the bacterial spreading map in this ward showed that bacterial densities were measured at the measuring points in the western part of the ward, which is located at the entrance to the ward, was more than the eastern parts (end of the ward). Figure 7 shows the possible spread of bacteria in some parts of the hospital.
According to the maps obtained from the spreading of bacteria in the intensive care unit (Fig. 7), the density of these microorganisms in the measured points located in the western part of the ward (entrance part) was more than the eastern parts (end of the ward). Due to the proximity of the entrance of the two infectious diseases and special care, the high presence of particles and bacteria in this area can be due to high traffic and lack of natural and artificial ventilation in this place.
The fungal spread distribution map in the intensive care unit shows that the rate of spread and dispersion of fungal aerosols is higher in the measuring points located in the western part and flow of pollutants has decreased while moving to the central and eastern (end of the ward). This situation indicates the concentration of fungi in the western part (entrance) of the intensive care unit. As mentioned above, the spread of bacteria has been higher in the intensive care unit in the western part. The following figure (Fig. 8) shows the possible spread of fungi in some parts of the hospital.
One of the reasons for the high presence of pollutants at the entrance of the ICU can be the lack of proper and adequate ventilation in this area, as well as the high traffic of the population and patients. On the other hand, in multi-bedroom wards of the hospital, including the intensive care unit, because there is a risk of infection of neighboring patients, reasonable measures should be taken to control the infection. In a study by Ching et al. carried out on reducing the risk of airborne infections in hospitals using hospital curtains, hospital curtains were called as physical barriers to disease transmission is potentially a simple but effective way to reduce the risk of infection; in this study, the effectiveness of hospital curtains in preventing the transmission of airborne diseases in hospital rooms has been investigated using numerical modeling. Among all case studies, it was found that the use of curtains between two beds can reduce the peak contaminant concentration for each neighboring patient in a bioaerosol dispersion process by up to 65% (Ching et al. 2008).