Microsporidiosis

John E. Bennett MD , in Mandell, Douglas, and Bennett's Principles and Practice of Infectious Diseases , 2020

Epidemiology

Microsporidian spores are commonly found in surface water, and human pathogenic microsporidia have been found in municipal water supplies, tertiary sewage effluent, and ground water. 113 – 118 It is likely that many of the Microsporidia are waterborne pathogens, and they can be transmitted by food. 119 Water contact has been found to be an independent risk factor for microsporidiosis in some studies 120,121 but not in others. 122,123 Outbreaks ofV. corneae infection have been associated with hot springs exposure 124 and exposure to soil. 125 , 126 E. cuniculi spores remain viable for 6 days when in water and 4 weeks when dry at 22°C, andNosema bombycis spores may remain viable for 10 years in distilled water. 127 Spores may be killed, however, by exposure for 30 minutes to 70% ethanol, 1% formaldehyde, or 2% Lysol or by autoclaving at 120°C for 10 minutes. 128 Most microsporidian infections are transmitted by oral ingestion of spores, with the site of initial infection being the GI tract. Viable infective spores of microsporidia are present in a number of body fluids (e.g., stool, urine, respiratory secretions) during infection, suggesting that person-to-person transmission can occur and that ocular infection may be transmitted by external autoinoculation caused by contaminated fingers. 129 It has been possible to transmitE. cuniculi via rectal infection in rabbits, suggesting the possibility of sexual transmission. 130 E. hellem has been demonstrated in the respiratory mucosa and in the prostate and urogenital tract of patients, raising the possibility of respiratory and sexual transmission in humans. 131,132 Person-to-person transmission is supported by concurrent infections in cohabiting homosexual men. 133 Although congenital transmission ofE. cuniculi has been demonstrated in rabbits, mice, dogs, horses, alpaca, foxes, and squirrel monkeys, no such congenital transmission has been demonstrated in humans. 134

There have been two clusters of microsporidiosis due to transplanted organs. 135 Three patients were infected by transplanted organs (kidney, kidney, and bilateral lungs) from anE. cuniculi–seropositive donor, and kidney biopsy provided the diagnosis in all three cases. Onset of symptoms (fever, renal dysfunction, and encephalopathy) was 7 to 10 weeks posttransplant, and death in one recipient of a kidney may have been directly related to failure of the transplanted organ. The surviving kidney recipient received 6 months of albendazole therapy and remained healthy afterward. 135 In the second cluster, which also involved three patients infected by transplanted organs (kidney, liver, heart/kidney), the patients presented with neurologic symptoms, including headache and encephalitis. 136 Kidney or urine specimens demonstratedE. cuniculi in these patients. The liver transplant patient responded to albendazole treatment. 136

Toxic effects of caustics (corrosives)

P.K. Gupta , in Fundamentals of Toxicology, 2016

23.3.2.1 Carbolic acid (phenol)

Carbolic acid is a poison that can be identified by its smell, which is commonly referred to as a phenolic odor or hospital odor. Pure phenol has a colorless, short, prismatic, needle-shaped, crystalline form. On exposure to air, it turns pink and liquefies. It is fat-soluble; therefore, it can attack the nervous system. It is also soluble in glycerin, ether, and alcohol, and it is slightly soluble in water. It is known specifically for its antiseptic or disinfectant property.

Other members of phenol group : Phenol has several derivates, namely, cresol, creosote, lysol, and dettol. These are absorbed orally, through intact skin, by the GI tract, through inhalation by the respiratory tract, per rectum, and per vaginum. The toxicological actions of these compounds are similar to phenol but less severe.

•

Cresol is a methyl phenol with meta, ortho, and para isomers. It is used as a disinfectant and antiseptic.

•

Creosote is a mixture of phenols consisting mainly of cresol and guiacol. It is used as a household remedy for coughs and is found in many proprietary preparations.

•

Resorcinol is a colorless crystalline substance used for the treatment of various skin diseases, including ringworm, psoriasis, and eczema.

•

Lysol is a 50% solution of cresol (3-methyl phenol) in saponified vegetable oil.

•

Thymol is an alkyl derivative of phenol obtained from volatile oils of Thymus vulgaris, Monarda punctata, or Trachyspermum ammi. It occurs in colorless crystals with a characteristic pungent odor and taste. Previously, it was used as an antihelminthic (for ankylostomiasis), antifungal, and antiseptic.

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Lice (Pediculosis)

John E. Bennett MD , in Mandell, Douglas, and Bennett's Principles and Practice of Infectious Diseases , 2020

Prevention

Prevention strategies for head lice include combinations of sanitizing the environment and, more important, eliminating all human reservoirs of carriage of head lice in households, apartments, housing complexes, homeless shelters, classrooms, and schools. Some common preventive interventions include the following: (1) avoiding contact with potentially contaminated items, such as hats, head sets, clothing, towels, combs, brushes, bedding, and upholstery; (2) soaking all combs and brushes in isopropyl alcohol or 2% Lysol solution; (3) sanitizing the household environment by hot-cycle washing and drying of all bedding, clothing, and headgear; and (4) inspecting high-risk schoolchildren for active head lice, viable nits, and nymphs. Lebwohl and colleagues 8 have recommended that "no-nit policies" in schools be abandoned. As noted, nonviable nits on hair shafts are simply empty egg cases and do not indicate active louse infestation. Dermoscopy can now clearly distinguish viable nits from hatched, empty nits and pseudonits immediately and reliably and offers a more sensitive screening tool for head lice infestations than inspection alone. 19

Prevention and control strategies for pediculosis corporis should include the following:

1

Hot-cycle washing and drying of all clothing and bedding

2

Clothing and body delousing with 1% permethrin dusting powder, especially in outbreak situations with potential for bacterial disease transmission

3

Institution of basic personal hygiene and sanitation measures, including showering, body washing, and clean clothing changes

Prevention strategies for pubic lice are similar to the prevention strategies for body lice and should include the following:

1

Hot-cycle washing and drying of all clothing and bedding

2

Institution of basic personal hygiene and sanitation measures

3

Treatment of sexual contacts with active infestations

4

Examination and laboratory testing of patients and their sexual contacts for other sexually transmitted diseases, especially crusted scabies and AIDS.

Medicinal Uses of Vinegar1

Carol S. Johnston , in Complementary and Alternative Therapies and the Aging Population, 2009

Antimicrobial Properties

Recent scientific investigations clearly demonstrate the antimicrobial properties of vinegar, but mainly in the context of food preparation as discussed above. Experts advise against using vinegar preparations as household disinfectants since they are less effective at inhibiting bacterial growth when compared to commercial disinfectants such as Lysol, Mr. Clean, and Clorox [13]. However, undiluted vinegar may be used effectively for cleaning dentures, and, unlike bleach solutions, vinegar residues left on dentures were not associated with mucosal damage [14]. In the popular media, vinegar is commonly recommended for treating nail fungus, head lice, and warts, yet scientific support for these treatment strategies is lacking. Although investigations have demonstrated the effectiveness of diluted vinegar (2% acetic acid solution at pH 2) for the treatment of ear infections (otitis externa, otitis media, and granular myringitis), the low pH of these solutions may irritate inflamed skin and damage cochlear outer hair cells [15].

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Shigellosis

Lee Goldman MD , in Goldman-Cecil Medicine , 2020

Prevention

Personal hygiene (in particular hand washing after handling the diapers of infected children and before food preparation), the sanitary disposal of feces, and protection of food and water sources from microbial contamination are essential to limit the spread of shigellosis. Preventing spread in daycare settings is a particular problem because it is so difficult to stop young children from constantly exploring their world, picking up bacteria on their hands, and bringing their fingers or contaminated fomites to their mouths. In such settings, it is particularly important for the adult caretakers to observe good personal hygiene and to supervise children in handwashing. Soap and water are sufficient, but hand sanitizers do work and may be more convenient. In environments where soap is unavailable, water used with sand or ash for scrubbing is helpful. Keeping infected children away from daycare until their stool is negative, if indeed cultures were done, or separating recently ill children from the susceptible has been recommended but is costly and not easy to accomplish. Household hygiene in the setting of an index case, including frequent handwashing, caution in the disposal of soiled diapers or underwear, regularly wiping down the area where these are collected with a disinfectant such as Lysol, and precautions in food preparation, can help limit intrahousehold spread. All of these measures are more difficult to implement in resource-limited environments without access to water for hygiene, let alone safe water for drinking or cooking, toilets or properly maintained latrines, ability to purchase disinfectants, or refrigeration to store food.

Vaccines, particularly for the more virulent species, would be useful. However, despite much effort and progress, a safe and effective vaccine has not been developed or approved for general use. 21 Vaccines againstShigella have been challenging to design because of the multiple species and serotypes, insufficient understanding of immunity and cross-protection, lack of a faithful animal model, and low commercial interest. In the United States populations for whom aShigella vaccine could be recommended include travelers or military personnel who are deployed to high-risk locations. In low-income countries, a multivalent broadly protective vaccine againstS. flexneri andS. sonnei, the two most commonShigella species in the last 20 years, would be extremely useful because shigellosis continues to account for a significant portion of the annual mortality due to diarrheal diseases. Our current understanding of immunity in shigellosis indicates that serotype-specific immunity is most important. Attenuated live oral vaccine candidates are probably most useful for travelers and military personnel deployed to high-risk areas; unfortunately, vaccines most effective in inducing immunity have also been the most reactogenic, causing fever and often mild diarrhea in recipients. For children in endemic areas current vaccines under clinical development are killed whole cells, 22 lipopolysaccharide (LPS) O antigen–protein conjugates, and subunit candidates. Newer-generation conjugate vaccines against LPS seem the most promising. Unless common protective antigens are identified, the most likely vaccine strategy will be based on a combination of serotype-specific antigens from the most prevalent and cross-protectiveShigella isolates. Refinement ofShigella human challenge models offer promise for acceleratingShigella vaccine development.

Animal Models of Tuberculosis

Devyani Dube , ... Suresh P. Vyas , in Animal Biotechnology, 2014

Protocols

This section of the chapter deals with describing the established protocols for infected animal models of TB, subsequent processing for analysis, and studying various aspects of biosafety that must be observed. All the protocols are illustrated by means of flow charts.

Preparing M. tuberculosis Inoculum for Aerosol Exposure

From the known concentration of M. tuberculosis stock solution, dilutions are prepared to infect mice or pigs by aerosols. To verify the amount of inoculums used in the nebulizer, a diluted aliquot of bacterial suspension is plated on 7H11 agar. Care should be taken during serial dilutions to prevent the generation of aerosol. All pipette tips, spills or drops should be disinfected with 5 % Lysol ( Flow Chart 2.1).

FLOW CHART 2.1.

Aerosol Infection of Mice Using the Middlebrook Apparatus

The Middlebrook airborne infection apparatus is the most widely used aerosol generation device. It is used to establish an animal model of pulmonary tuberculosis to resemble normal route and site of infection in humans. This instrument consists of an aerosol chamber that contains a basket or cage with five compartments that can accommodate 25 mice. Compressed air flows through the nebulizer and produces a fine mist of bacterial suspension carried into the aerosol chamber, which is inhaled by the animals. Finally, a UV lamp is used to decontaminate the surface (Flow Chart 2.2, Figure 2.2).

FLOW CHART 2.2.

FIGURE 2.2. Glass-Col apparatus. This instrument is the advancement of earlier models developed by Middlebrook. 80 to 100 mice can be exposed to aerosol at one time.

Aerosol Infection of Guinea Pigs Using a Madison Chamber

A Madison chamber is an aerosol generation device used for guinea pigs because of their larger size. (Figures 2.3 and 2.4 ) The surfaces of the Biosafety Class II Cabinet should be sterilized with 5% Lysol and 70% ethanol.

FIGURE 2.3. (A) Collison nebulizer unit, complete with the surrounding glass nebulizer jar, for the Madison guinea pig aerosol chamber, (B) Photohelic meter for Madison chamber.

FIGURE 2.4. The exterior (A) and interior (B) of the Madison chamber control box.

A test run to ensure adequate pressure driving is shown in Flow Chart 2.3.

FLOW CHART 2.3.

Bacterial Loading

Before bacterial loading, each of the corresponding guinea pig cages should be labeled by writing "aerosolized with M. tuberculosis on date/initials." A stainless steel container filled with 5% Lysol solution should also be placed next to the infection chamber ( Flow Chart 2.4).

FLOW CHART 2.4.

Intravenous Infection of Mice with M. Tuberculosis

Intravenous infection requires absolute concentration, and good eyesight with no distraction. The mouse is placed in a restraint device so that the tail can be immobilized and then the injectate is injected into the lateral vein (Flow Chart 2.5).

FLOW CHART 2.5.

Isolation of Samples for Determining M. tuberculosis Load by RT-PCR

Expression of the multiple genes during infection of M. tuberculosis can be measured by isolation of total RNA and then running a PCR assay. During the PCR assay, contamination of test samples with RNAase and extraneous sources of DNA should be prevented. Samples containing RNA and cDNA should be dealt in separate working areas. The protocol describes sample collection from the infected tissues to be processed for the PCR analysis (Flow Chart 2.6).

FLOW CHART 2.6.

Determination of Bacterial Loads in Target Organs

Processing of infected animals should be done in a biosafety cabinet. All the necessary items should be kept in the biosafety hood without restricting the airflow. Animals should be euthanized according to the guidelines provided by the IACUC. Due to the chronic nature of the infection, the design of the study depends on the time course to be studied. Number of animals per time point should also be considered, which can be determined through statistical power calculation. For the bacterial load curve, a power of > 0.8 can be achieved using four to five mice (Flow Chart 2.7).

FLOW CHART 2.7.

The process includes:

1.

Preparation of agar plates

2.

Bacterial count set-up

3.

Necropsy

4.

Homogenization of tissues

5.

Plating

6.

Bacterial count colonies

Preparation of Lungs or Other Tissues for Histology

Useful information about the host response can be acquired by the histological examination of organs from infected animals. The size of granulomas, their make-up in terms of lymphocytes and macrophages, the degree of lung tissue they are consolidating, the development of necrosis, and so forth, can be invaluable information. It is important that the specimens are frozen as soon as they are collected to preserve their morphology and integrity of the antigens. Sections are stained with hematoxylin and eosin after fixation and paraffin embedding (Flow Chart 2.8).

FLOW CHART 2.8.

Preparation of Lung Cell Suspension

Cells suspensions from naïve and M. tuberculosis infected mice are used for single cell analysis studies by flow cytometry by cell culture. Cells from the culture of infected animals provide information such as cell activation, proliferation and cytokines as well as chemokine production (Flow Chart 2.9).

FLOW CHART 2.9.

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Dish and Household Cleaning

Gregory Szewczyk , Karen Wisniewski , in Handbook for Cleaning/Decontamination of Surfaces, 2007

1.2. Sanitization in Household Cleaners

Traditional household cleaners, whether used on household surfaces or on dishes, are primarily focused on basic cleaning rather than with sanitization. For the last thirty years, products that both clean and kill germs have been popular in the developed world in bathroom cleaning; until recently this type of bathroom cleaner constituted the largest group of germ killing cleaners. However, the acceptance of germ killing cleaners was slower in other parts of the house. Except for specific uses such as garbage pail cleaning or wiping up surfaces in infant rooms, germ killing cleaners were limited to the bathroom. With a few exceptions, hand and surface sanitizing cleaners for hand dishwashing are not prevalent. However, dishwashing liquids that double as antibacterial hand soaps are prevalent in the United States. Germ killing tends to be a value added benefit to a product whose primary purpose is soil removal.

Following the confirmation of the germ theory of disease, physicians began using disinfecting agents in hospitals and other medical situations. This started the spread of disinfectants for domestic uses. Household disinfection goes back to the late 1800s when products like Lysol ® household disinfectant were first sold. These early disinfectant products were not cleaners and usually contained phenol, an ingredient seldom used today because of toxicity concerns. (The original Lysol® concentrate still uses phenol.) It must also be remembered that around the turn of the twentieth century, there were outbreaks of dangerous infectious diseases like typhoid and diphtheria commonplace in America and Europe, especially in crowded cities. Consumers needed education, to replace the "miasma" theory of disease with germ theory which happened gradually from the 1880s into the 1920s [2]. Manufacturers of household disinfectants were part of this education about germs around the household, with their products sold as a way of making the home healthier in addition to cleaning. The consumer understanding of such disinfecting products was "a drug item rather than a household helper". This consumer attitude persisted into the 1960s, when housewives felt that to be a Lysol® user they had to "just have come through a flood or hurricane or be in the middle of an epidemic" [3]. This severely limited the use of disinfectants in the home. By the middle of the twentieth century, many women were working outside the home and no longer had the time (or the inclination) to do two step process of cleaning followed by disinfection. These two forces resulted in the formulation of products that disinfected as well as cleaned, an effort that started in the late 1940s [4]. It can be argued whether such household cleaners are still so necessary in developed nations without widespread outbreaks of serious contagious diseases. Despite this, anti-bacterial and disinfectant cleaners continue to have a significant presence in the marketplace, especially in North America. Indeed, the number of types of cleaners claiming some degree of germ kill has increased over recent years. The trend started with disinfecting all purpose cleaners, toilet bowl, and bathroom cleaners and evolved into an assortment that includes disinfecting glass and surface sprays, toilet and general purpose wipes, and hand washes. It may be argued that with the concerns over SARS, avian flu, and other such diseases, as well as worries over salmonella contaminated chicken, Escherichia coli tainted hamburger, and germy kitchen sponges, these products probably will increase in popularity.

Disinfecting products are often government regulated, with agencies specifying the testing methods, labeling regulations, and product claims. Germ kill claims are not visible to consumers like cleaning claims so they cannot self-assess product efficacy. This makes regulation and consumer advocacy important for disinfecting products. More detail about the regulations for household cleaners will be discussed when test methods for the household products are discussed.

A note on nomenclature is important for products labeled as "sanitizers" and "disinfectants". Although these terms are almost interchangeable in many consumer's minds, they are not identical to regulatory agencies. The EPA is the Environmental Protection Agency of the United States government which regulates consumer products that claim to kill microorganisms on non-living surfaces. To the EPA these terms constitute different levels of antimicrobial control [5]. A sanitizer does not necessarily eliminate microorganisms but instead reduces them to "a level considered safe". A disinfectant destroys or irreversibly inactivates infectious fungi and bacteria (but not their spores) on hard surfaces. A sterilizer destroys or eliminates all forms of microbial life. These distinctions in the levels of organism reduction are lost on most consumers. Most products in the developed world are tested at the disinfectant level, but some make label claims that invoke sanitization ("kills 99.9% germs"). A special case is that of food contact surfaces where a sanitizer is a 5 log reduction of organisms in 30 s. Another term used in the developed world is "antibacterial". Products labeled with this term usually have "disinfectant" in smaller type somewhere else on the bottle. Disinfectant is the older label term and sometimes perceived as the more "medicinal" or "strong" term. "Disinfectant" is in large type (usually) on bathroom cleaners, while kitchen use products usually use the "antibacterial" term. (From a textbook perspective, "antibacterial" indicates products that try to control microbes, whether it is to kill them (bactericidal) or just limit the growth (bacteriostatic).) There was evidently such a strong association between Lysol® or bathroom products and disinfectant that this term was viewed as inappropriate for use in the kitchen on food preparation or dish surfaces. The first antibacterial kitchen product launched was an antibacterial hand dishwashing liquid. (Since these products' antibacterial claims are on hands, they are regulated in the United States by the Food and Drug Administration; the EPA regulates products whose claims are for germ killing on inanimate surfaces. Other general kitchen surface cleaners, usually as sprays or wipes, followed using the antibacterial term pioneered by the dishwashing product rather than the disinfectant term used by bathroom cleaners. The actual chemical mechanism of kill, the technical performance and the regulation of the products are identical whether they say "disinfectant" or "antibacterial" as long as they are used on the same type of surfaces. This is evidenced by the "disinfection" subtitling on antibacterial products in the United States used to clean hard surfaces. The terms on the labels are mostly due to consumer sensibilities rather than technical differences, and the consumers seem confused over what they mean [6].

Disinfecting household cleaners are a highly regulated category of consumer products in developed nations. It is incumbent on the formulator to be familiar with the regulations of the country of sale. There are two key systems of regulation shown in North America by the Canadian and US procedures. Under a "monograph" system, such as that followed in Canada, a published list of rules determines which products may be labeled as disinfectants. If a formula follows the rules including a list of approved active ingredients, a mandated effective concentration (in use) for each ingredient, and the types of end-use products, it can carry the disinfectant language. The alternate system is more rigorous and uses exhaustive testing at each level of formulation. Under the EPA system, for example, the disinfectant component must first be tested alone for efficacy, and if effective, be registered with the regulating agency as an approved active ingredient. Then the active ingredient must be tested in the proposed formulation to assure that it retains its efficacy in both fresh and aged product. Toxicological and chemical properties of both the active and end use formula are also scrutinized [7]. This specific formula, including the exact aesthetic (color and perfume) and inert ingredients, is then registered as an approved formula. The monograph system has the advantage of making the registration process quick and relatively easy, but getting new ingredients added to the monograph requires laborious petitioning to change the monograph. A monograph system limits formulation creativity and opportunities for exclusivity. The testing system allows for multiplicity of disinfectant ingredients and creative formulation. However, it is time and resource intensive to perform all the mandated tests under all the specified conditions. A US EPA formula registration can take one to five years.

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Immunology of Infection

Diane J. Ordway , lan M. Orme , in Methods in Microbiology, 2010

F Analysis of Cell Surface Expression of Markers in the Mouse by Flow Cytometry

To analyze the expression of surface markers on cells derived from tissues, cells are prepared as above and re-suspended at a concentration of 5   ×   106 cells/ml in PBS containing 0.1% sodium azide. Cells are incubated in PBS containing 0.1% sodium azide on ice for at least 30   min. This incubation is an important step if one is analyzing the expression of cell surface activation molecules; this is because it is our experience that the antibody-mediated ligation of activation markers on the cell surface results in the up-regulation of several molecules that are used to determine cell activation status. For example, immediate incubation of cells with anti-CD44 and anti-CD45RB results in the cells becoming 'blast cells' (as determined by an increase in scatter by flow cytometer analysis) and in the increased expression of CD44 on the cell surface.

Once cells have been fixed, they are dispensed into 96-well plates (200 μl per well) and centrifuged to pellet the cells. The supernatant is removed by gently inverting the plate and discarding the contents into a vessel containing a 5% Lysol solution. Antibody is added to the cells at appropriate concentrations, and the plate incubated for 30  min at 4°   C in the dark. Following two washes in d-RPMI the cells are analyzed on a flow cytometer.

Recommended stains include the following:

Discriminating between cell populations: FITC anti-CD19 or CD45R/B220 (B cells), PE anti-CD3e (T ells), PerCP anti-CD4 (T-cell subset), APC anti-CD8 (T-cell subset) or FITC anti-NKl.1 (NK cell), PE anti-CD3e (T cell), PerCP anti-CD4 (T-cell subset), APC anti-CD8 (T-cell subset).

Effector/memory T-cell differentiation: FITC anti-CD44 (T-cell activation), PE anti-CD62L (T-cell memory), PerCP anti-CD3e (T cell), APC anti-CD4 or CD8 (T-cell subset) or FITC anti-CD44 (T-cell activation), PE anti-CD62L (T-cell memory), PerCP anti-CCR7 (T-cell memory), APC anti-CD4 or CD8 (T-cell subset).

Regulatory T-cell evaluation: FITC anti-CD25 (T-cell activation), PE anti-Foxp3 (Treg marker), PerCP anti-CD3e (T cell), APC anti-CD4 or CD8 (T-cell subset).

Macrophage/DC evaluation: FITC anti-CD80 (cell activation), PE anti-MHC class II (cell activation), PerCP anti-CD11b (macrophage/DC differentiation), APC anti-C11c (macrophage/DC differentiation).

It is essential to include isotype controls for each individual antibody isotype that is used, in order to appreciate the contribution of non-specific binding.

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Biocides

David R. Karsa , in Handbook for Cleaning/Decontamination of Surfaces, 2007

3.12 Phenolics

Phenolic biocides have been used since the surgeon, Lister, introduced phenol into surgical practice in 1867. Phenolic biocides are used across a wide range of applications including disinfectants, antiseptics, surgical scrubs, toilet soaps, cosmetics, etc. with the exception of food contact application where taint is a problem. Phenolic derivatives are more active in the relatively insoluble acidic form against bacteria and fungi, but their effectiveness against viruses and spores is not particularly good. They are also deactivated by organic soils.

Traditionally, phenols have been solubilised into a usable form using soaps. Simple soaps, such as potassium laurate, are susceptible to hard water and hence synthetic detergents are often used such as sulphonated castor oil, alkylbenzene sulphonates or alkylether sulphates. Here, careful formulation is required. An equilibrium exists between free phenolic biocide in solution and that in micelles. Some surfactants can "over-solubilise" the phenolic biocide reducing its effective availability.

The early coal tar disinfectants were introduced in the 1880s based on three types of product from coal tar distillates. These were used to produce the so-called, "clear fluids", "black fluids" and "white fluids". Clear fluids were based on derivatives such as cresols (Lysol 1897) and xylenols (Sudol 1953) solubilised in soap or surfactant to give a clear aqueous dilution. Black fluids (1887) contain a large number of phenolic derivatives which distil over as high boiling tar acids (250–310°C). These include tri- and tetra-methylphenols, propyl/butyl phenols, methyl resorcinols and naphthols plus neutral hydrocarbon tar oils. Again these may be solubilised with soaps or surfactants to form clear black liquids which readily form emulsions when added to water. They are effective under heavy soiling conditions against both bacteria and fungi. White fluids also contain high boiling tar acids, but this time they are formulated into emulsion concentrates by soap and emulsion stabilisers, such as casein, xathan gums, etc. They are also effective in conditions of heavy soiling.

Substitution of an alkyl group of up to six carbon atoms into the phenol ring (preferably in the para-position) increases the biocidal activity, probably by increasing surface activity. Halogenation also increases the anti-bactericidal activity of a phenol, for example the trichorphenols which are popular antiseptics and effective fungicides. However, some products, such as the wood preservative pentachlorophenol, have been banned in Europe due to their persistence in the environment and detrimental effect on sewage treatment bacteria.

A combination of alkyl and halogen substitution into the phenol confers the greatest antibacterial activity, when the alkyl group is ortho- to the phenol group and the halogen is para- to it.

Bis-phenols, compounds containing two phenyl groups, have been developed as commercial biocides. The two phenol groups may be connected directly or separated by a methylene group or an oxygen or sulphur atom. Examples are dichlorophen and triclosan (Figure F.2.22).

Figure F.2.22. Dichlorophen and triclosan

Dichlorophen has been used as a preservative for toiletries, textiles and cutting oils and to prevent bacterial growth in water-cooling systems. Triclosan is widely used as a preservative in many formulated products. It is also used in handcleaning gels and medicated soaps. Other products, such as 2-phenylphenol, are effective fungicides and bactericides, sometimes used in pine-type disinfectants.

Again successful formulation of this large range of phenolics requires some skill and understanding, and careful consideration has to be given to pH, the choice of soap or surfactant solubiliser and deactivation by their ability to form inactive complexes with nonionic surfactants.

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Post-processing

John Cyganowski , Herb Lutz , in Ultrafiltration for Bioprocessing, 2015

8.4 Post CIP measurements

8.4.1 Residuals

Removal of cleaners and sanitizers by flushing is needed to ensure no carryover into the next batch processing. Carryover would degrade the product and cause quality issues as well as fouling the membrane. Simple CIP agents like NaOH, peracetic acid and NaOCl are easily measured by pH, conductivity, or chlorine assay kits (designed for environmental purposes or recreational pool maintenance), respectively. Verifying the removal of more complex CIP preparations (detergents, enzymes, chelating agents, etc.) will require more complex assays or even specific assay development. Consult your CIP chemical vendor for recommendations. Residual total organic carbon (TOC) measurement is a sensitive reliable method to assess organic carbon levels in the flush out streams from the membranes and system. This testing can provide direct evidence of the effectiveness and consistency of the cleaning cycle.

A key consideration for storage agents is the ability to validate the removal of the agent. There is a test-strip-based colorimetric assay for alkyldiethylbenzylammonium chloride (Lysol Brand Sanitizer) and peracetic acid with 1 ppm sensitivity. A simple conductivity meter can be used to demonstrate removal of the other sanitizing agents in the following list ( Table 8.10).

8.4.2 Normalized water permeability

The normalized water permeability (NWP) test is a recommended method to assess the effectiveness of the CIP process to restore process flux rates. NWP uses water permeability (LMH/psi) as a measure of membrane cleanliness:

(8.4) NWP = ( litres per hour volumetric permeate flow ) ( TCF ) ( TMP psid ) ( membrane area m 2 )

TMP = ((P feed + P retentate) / 2 – P permeate) as the module average transmembrane pressure. TCF = (viscosity of water at measured temperature)/(viscosity of water @ 25°C) as temperature correction factor (see Appendix).

As described in Section 8.4, the hydraulics of modules are complex and the calculated NWP may be affected by membrane permeability and the selection of feed flow and TMP operating conditions. The calculated NWP value is subject to pressure gauge measurement error so it is useful to run at least at 10 psi TMP to minimize this effect. For high-permeability membranes, low feed flow and high TMP will cause dead-end flow where all the feed goes into the permeate stream. High permeate flows can also cause a pressure drop in the permeate stream so the TMP calculation in equation (8.4) will be thrown off. Air bubbles can build up in the retentate and blind off the membrane to alter the measured NWP. It is critical that the retentate flow is at least 10% of the feed flow to avoid this. This can be achieved through higher feed flows. For lower permeability membranes, there is no need to run at high feed flows as this can accentuate feed channel pressure drop effects.

NWP is measured for a new set of membranes at installation (Section 8.6), after flush and after the first CIP cycle. The first CIP removes membrane preservatives and conditions the membrane prior to the first exposure to process fluid. This measurement becomes the baseline, which is compared to subsequent NWP measurements taken post-CIP. The acceptance criterion for cleaning efficiency is membrane and application specific and may vary between plants. A typical criterion for regenerated cellulose NWP is that if the membrane NWP is ±20% of the baseline NWP (after the initial cleaning), process reproducibility will result. 6 For polyethersulfone membranes and/or fouling feeds, larger variations in NWP of ±40% are seen. Consistency in the NWP measurement rather than an absolute limit is the best indicator of successful cleaning. For example, suppose that for a particular protein, a 40% decrease in NWP is observed upon first use but that subsequent NWP measurements were within ±20% of the reduced NWP. A consistent NWP coupled with consistent process fluxes (process cycle times), consistent yields, acceptable product quality, and passing system integrity tests, would lead one to conclude that the CIP process was efficiently cleaning the membranes and maintaining system performance. NWP is an important indicator of cleaning efficiency, but should be evaluated within the context of process performance.

Application of NWP as a measure of cleaning effectiveness involves tracking NWP from run-to-run. An initial value should be measured on modules that have been initially cleaned but have not seen protein. It has been observed that NWP declines during the first cycle of operation, even without exposure to protein. This is attributed to the cassette screen compressing into the membrane and blinding off some of the membrane area. There is some variability in the measurement of the NWP. For example, it can change with the operating conditions. NWP should be measured under consistent conditions for each run to minimize variability. If the NWP drops by more than 20%, it is possible that the cleaning cycle was not effective and an investigation should be made. If the cleaning agent was incorrectly formulated or lost some of its potency due to age, this could be a cause of the decline. Provision should be made for a re-cleaning option. In addition, if the NWP steadily decreases from run-to-run, this is a sign of inadequate cleaning. It would appear that some component is building up on the membrane and not being removed by the cleaning cycle.

Post-use integrity testing is generally not performed for an ultrafiltration system. Pre-use testing is generally used (Section 6.3). That is because defects are generally introduced with the first time module installation in large systems. After operation, the modules are not disturbed and the integrity is preserved. One exception occurs when modules are subjected to thermal cycles during the cleaning process and system compression is fixed at a given position. Cycling may reduce compression and cause sealing defects. Rechecking compression or the use of hydraulic cups providing a constant load can remedy this. Before extended storage of modules in the holder of 7 days or more, it may be useful to run an integrity test.

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